GenPlay, Einstein Genome Analyzer - User contributions [en]

Documentation

2013-10-11T20:13:32Z

Nicolas: /* The Welcome screen */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name.
[[image:mg_basics_project name.png|center|frame|Text field to define the project name]]

The precision of the project will change the number of bits used to code numbers.
* High-Precision: Numbers are coded using 32 bits which offers the highest precision level in GenPlay.
* Low-Precision: Numbers are coded using 16 bits. It may be useful to lower memory usage. However, the maximum score is 65504 and decimals may be rounded in a different way ([http://en.wikipedia.org/wiki/Half-precision_floating-point_format here] for more information).
[[image:precision.png|center|frame|Project precision]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly.
[[image:mg_basics_assembly_chooser.png|center|frame|Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser, users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in figure below.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Empty welcome screen for multi-genome project]]
===== Single Genome Project =====
The Single Genome Project is the most common/regular project in GenPlay. If you do not know or understand yet what the Multi Genome Project is, please use the Single Genome Project.
===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
[[image:load_project.png|right|thumb|150px|Load an existing project]]
In order to load a project, the user has to select the "Load an existing project" option. 
The list of the 5 last projects shows on the lower part of the dialog. An additional option "Other" will let the user select a GenPlay project file to load. 
The upper part updates automatically when selecting a project in order to remind the following information:
* Name: The name of the project.
* Precision: The precision of the project, either high or low.
* Genome: The genome used.
* Project type: The type of project, either single or multi-genome.
* Last modified: The last time the project has been modified.
* Track number: The number of track in the project.
 

== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#MGFiltersExplanation|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
<div id="MGFiltersExplanation"></div>
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>

==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

File:Precision.png

2013-10-11T20:00:19Z

Nicolas:

File:Mg basics empty welcome screen.png

2013-10-11T19:55:57Z

Nicolas: uploaded a new version of "File:Mg basics empty welcome screen.png"

Documentation

2013-10-11T19:52:43Z

Nicolas: /* Load Project */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
[[image:load_project.png|right|thumb|150px|Load an existing project]]
In order to load a project, the user has to select the "Load an existing project" option. 
The list of the 5 last projects shows on the lower part of the dialog. An additional option "Other" will let the user select a GenPlay project file to load. 
The upper part updates automatically when selecting a project in order to remind the following information:
* Name: The name of the project.
* Precision: The precision of the project, either high or low.
* Genome: The genome used.
* Project type: The type of project, either single or multi-genome.
* Last modified: The last time the project has been modified.
* Track number: The number of track in the project.
 

== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#MGFiltersExplanation|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
<div id="MGFiltersExplanation"></div>
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>

==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

Documentation

2013-10-11T19:52:20Z

Nicolas: /* Load Project */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
[[image:load_project.png|right|thumb|200px|Load an existing project]]
In order to load a project, the user has to select the "Load an existing project" option. 
The list of the 5 last projects shows on the lower part of the dialog. An additional option "Other" will let the user select a GenPlay project file to load. 
The upper part updates automatically when selecting a project in order to remind the following information:
* Name: The name of the project.
* Precision: The precision of the project, either high or low.
* Genome: The genome used.
* Project type: The type of project, either single or multi-genome.
* Last modified: The last time the project has been modified.
* Track number: The number of track in the project.
 

== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#MGFiltersExplanation|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
<div id="MGFiltersExplanation"></div>
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>

==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

File:Load project.png

2013-10-11T19:44:43Z

Nicolas:

Documentation

2013-10-11T19:37:32Z

Nicolas: /* Filters */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#MGFiltersExplanation|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
<div id="MGFiltersExplanation"></div>
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>

==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

Documentation

2013-10-11T19:37:00Z

Nicolas: /* Filters */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#Variant Layer Operations:Filters|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
<div id="MGFiltersExplanation"></div>
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>

==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

Documentation

2013-10-11T19:35:33Z

Nicolas: /* Filters */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#Variant Layer Operations:Filters|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>
==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

Multi-Genome Tutorial

2013-10-11T19:32:28Z

Nicolas: /* Getting started */

== Getting started ==
In order to set up and manage a Multi-Genome Project in Genplay, please refer to the following sections of the documentation:
* [[Documentation#Multi Genome Project|Loading a Multi Genome Project]]
* [[Documentation#Loading a Variant Layer|Loading a Variant Layer]]
* [[Documentation#Variant Layer Operations|Variant Layer operations]]
 

== Conversion between NCBI36/hg18 and GRCh37/hg19 ==
=== Description ===
This tutorial will explain how to display at the same time tracks mapped on genome assembly NCBI36/hg18 or GRCh37/hg19. In the example, user will be able to see all the modifications on the NCBI36/hg18 genome leading to the GRCh37/hg19 reference genome.

=== Files ===
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_settings.xml XML settings file]
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_sv.vcf.gz VCF file]
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_sv.vcf.gz.tbi Indexed VCF file (Tabix)]
*[http://www.genplay.net/library/Human/hg19/Gene_Annotation/RefSeq_From_UCSC_04-23-10(hg19).bed Refseq BED file for GRCh37/hg19]
*[http://www.genplay.net/library/Human/hg18/Gene_Annotation/RefSeq_From_UCSC_04-23-10(hg18).bed Refseq BED file for NCBI36/hg18]

=== Steps ===
==== Project settings ====
===== Project name =====
User must choose a name for a new project; here the name is ''GenPlay-MG – Reference genome tutorial'' (Figure 1).
[[image:mg_hg18tohg19_project_name.png|center|frame|Figure 1: Project name]]

===== Project assembly =====
According to bed files provided in this tutorial, the reference genome is GRCh37/hg19. User has to select the ''mammal'' clade, the ''human'' genome and the ''Feb 2009 (GRCh37/hg19)'' assembly as in Figure 2.
[[image:mg_hg18tohg19_project_assembly.png|center|frame|Figure 2: Project assembly]]

===== Chromosome selection =====
The VCF file is about Structural Variants and contains information for chromosomes 1 to 22 and chromosomes X and Y. User can select to load on or more chromosomes by clicking on the settings button next to the assembly name (Figure 3).
[[image:mg_hg18tohg19_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

===== VCF Loading =====
'''''Manually''''' 
To load VCF files , users must first fill the column lists and then select from the list the appropriate data. The VCF Loader appears after clicking on the ''Edit'' button from the welcome screen. The bottom left part of the VCF Loader contains the ''Column list edition'' section. User has to select a column and click on ''Edit'' button in order to show the associated list.
Only one VCF file is going to be loaded for this tutorial. The VCF file contains differences between the reference genome NCBI36/hg18 and the reference genome GRCh37/hg19.
 

''Group'' column 
This tutorial compares reference genome; a generic group name can be ''Reference genome''.
On the ''Group name list editor'', user clicks on the plus button to show the input text box and fills it (Figure 4). 
The ''Group name list editor'' should looks like the Figure 5 below. 
Once the values has been added to the list, it can be saved by closing the "Group name list editor window" 
''value:'' ''' Reference genome'''
<gallery widths=350px perrow=2>
image:mg_hg18tohg19_group_input.png|Figure 4: Group name input dialog
image:mg_hg18tohg19_group_editor.png|Figure 5: Group name editor
</gallery>
 

''Genome'' column 
The genome name is an Alias for the selected raw name. In this tutorial, the genome name is going to be '''Hg18'''.
On the ''Genome name list editor'', user clicks on the plus button to invoke the input text box and fills it (Figure 6). 
The ''Genome name list editor'' should looks like the Figure 7 below. 
Once the values has been added to the list, it can be saved by closing the "Genome name list editor window" 
''value:'' '''Hg18'''
<gallery widths=350px perrow=2>
image:mg_hg18tohg19_genome_input.png|Figure 6: Genome name input dialog
image:mg_hg18tohg19_genome_editor.png|Figure 7: Genome name editor
</gallery>
 

''Type'' column 
This field cannot be edited by the users. The provided VCF file is a Structural Variant type, user therefore has to choose '''SV''' (Figure 8). 
''value:'' '''SV'''
[[image:mg_hg18tohg19_type.png|center|frame|Figure 8: VCF type list]]
 

''File'' column 
Once the VCF file is downloaded, user has to open the ''File list editor'', user clicks on the plus button to show the file chooser dialog and choose the VCF file according to its location. 
''value:'' '''VCF path'''
[[image:mg_hg18tohg19_file_editor.png|center|frame|Figure 9: VCF File editor]]
 

''Raw name(s)'' column 
The raw name list is automatically filled. In the case of this tutorial there is only one genome: '''NCBI36''' (Figure 10). 
''value:'' '''NCBI36'''
[[image:mg_hg18tohg19_raw_name.png|center|frame|Figure 10: Raw name list]]
Again, value is saved by closing the windows
 

'''''Import XML settings''''' 
In order to set the project with ease, user can import the settings using the XML file above. Please be careful about the VCF path, user must changes it directly on the xml file if he wants to use the import function.
 

'''''Conclusion''''' 
Finally, the screen should be like the one on Figure 11.
[[image:mg_hg18tohg19_vcf_loader.png|center|frame|Figure 11: VCF loader]]

===== Conclusion =====
The welcome screen should finally be similar to the Figure 12.
[[image:mg_hg18tohg19_welcome_screen.png|center|frame|Figure 12: Welcome screen]]

The "Create" button will create the project and will run the synchronization.

==== GRCh37/hg19 genes loading ====
To load a file, user has to do a right click on the left part of the track. Then to choose "Load Gene Track", a file chooser appears to select the file given in this tutorial. After having chosen the BED file, a new selection box appears (Figure 13).
[[image:mg_hg18tohg19_genome_selector_01.png|center|frame|Figure 13: Genome selection dialog for GRCh37/hg19 genes file]]

This box asks which genome is related to the BED file. Here, user has to choose "Feb 2009 (GFCh37/hg19)" option because the BED file contains information about that genome.
Gene file for GRCh37/hg19 reference has been loaded.

==== NCBI36/hg18 genes loading ====
The same operation as loading a gene files for GRCh37/hg19 reference genome. The only step changing is to choose the "Reference genome - hg18 (NCBI36)" option after the BED file selection (Figure 14)
[[image:mg_hg18tohg19_genome_selector_02.png|center|frame|Figure 14: Genome selection dialog for NCBI36/hg18 genes file]]

==== Conclusion ====
User can navigate into the different chromosomes and visualizes differences between both genomes using the stripes. All genes are perfectly synchronized and are display according to the meta-genome coordinates.

The Figure 15 shows an example of the result of this tutorial. It is possible to see deletions (in red) and insertions (in green) in the NCBI36/Hg18 reference genome compare to the GCh37/Hg19 reference genome. 
Chromosome: ''chr1'' 
Position: ''143,822,670'' 
[[image:mg_hg18tohg19_genplay_gene_example.png|center|thumb|800px|Figure 14: GenPlay-MG (chr1:143,822,670)]]

Multi-Genome Tutorial

2013-10-11T19:32:11Z

Nicolas:

== Getting started ==
In order to set up and manage a Multi-Genome Project in Genplay, please refer to the following sections of the documentation:
* [[Documentation#Multi Genome Project|Loading a Multi Genome Project]]
* [[Documentation#Loading a Variant Layer|Loading a Variant Layer]]
* [[Documentation#Variant Layer Operations|Variant Layer operations]]

== Conversion between NCBI36/hg18 and GRCh37/hg19 ==
=== Description ===
This tutorial will explain how to display at the same time tracks mapped on genome assembly NCBI36/hg18 or GRCh37/hg19. In the example, user will be able to see all the modifications on the NCBI36/hg18 genome leading to the GRCh37/hg19 reference genome.

=== Files ===
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_settings.xml XML settings file]
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_sv.vcf.gz VCF file]
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_sv.vcf.gz.tbi Indexed VCF file (Tabix)]
*[http://www.genplay.net/library/Human/hg19/Gene_Annotation/RefSeq_From_UCSC_04-23-10(hg19).bed Refseq BED file for GRCh37/hg19]
*[http://www.genplay.net/library/Human/hg18/Gene_Annotation/RefSeq_From_UCSC_04-23-10(hg18).bed Refseq BED file for NCBI36/hg18]

=== Steps ===
==== Project settings ====
===== Project name =====
User must choose a name for a new project; here the name is ''GenPlay-MG – Reference genome tutorial'' (Figure 1).
[[image:mg_hg18tohg19_project_name.png|center|frame|Figure 1: Project name]]

===== Project assembly =====
According to bed files provided in this tutorial, the reference genome is GRCh37/hg19. User has to select the ''mammal'' clade, the ''human'' genome and the ''Feb 2009 (GRCh37/hg19)'' assembly as in Figure 2.
[[image:mg_hg18tohg19_project_assembly.png|center|frame|Figure 2: Project assembly]]

===== Chromosome selection =====
The VCF file is about Structural Variants and contains information for chromosomes 1 to 22 and chromosomes X and Y. User can select to load on or more chromosomes by clicking on the settings button next to the assembly name (Figure 3).
[[image:mg_hg18tohg19_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

===== VCF Loading =====
'''''Manually''''' 
To load VCF files , users must first fill the column lists and then select from the list the appropriate data. The VCF Loader appears after clicking on the ''Edit'' button from the welcome screen. The bottom left part of the VCF Loader contains the ''Column list edition'' section. User has to select a column and click on ''Edit'' button in order to show the associated list.
Only one VCF file is going to be loaded for this tutorial. The VCF file contains differences between the reference genome NCBI36/hg18 and the reference genome GRCh37/hg19.
 

''Group'' column 
This tutorial compares reference genome; a generic group name can be ''Reference genome''.
On the ''Group name list editor'', user clicks on the plus button to show the input text box and fills it (Figure 4). 
The ''Group name list editor'' should looks like the Figure 5 below. 
Once the values has been added to the list, it can be saved by closing the "Group name list editor window" 
''value:'' ''' Reference genome'''
<gallery widths=350px perrow=2>
image:mg_hg18tohg19_group_input.png|Figure 4: Group name input dialog
image:mg_hg18tohg19_group_editor.png|Figure 5: Group name editor
</gallery>
 

''Genome'' column 
The genome name is an Alias for the selected raw name. In this tutorial, the genome name is going to be '''Hg18'''.
On the ''Genome name list editor'', user clicks on the plus button to invoke the input text box and fills it (Figure 6). 
The ''Genome name list editor'' should looks like the Figure 7 below. 
Once the values has been added to the list, it can be saved by closing the "Genome name list editor window" 
''value:'' '''Hg18'''
<gallery widths=350px perrow=2>
image:mg_hg18tohg19_genome_input.png|Figure 6: Genome name input dialog
image:mg_hg18tohg19_genome_editor.png|Figure 7: Genome name editor
</gallery>
 

''Type'' column 
This field cannot be edited by the users. The provided VCF file is a Structural Variant type, user therefore has to choose '''SV''' (Figure 8). 
''value:'' '''SV'''
[[image:mg_hg18tohg19_type.png|center|frame|Figure 8: VCF type list]]
 

''File'' column 
Once the VCF file is downloaded, user has to open the ''File list editor'', user clicks on the plus button to show the file chooser dialog and choose the VCF file according to its location. 
''value:'' '''VCF path'''
[[image:mg_hg18tohg19_file_editor.png|center|frame|Figure 9: VCF File editor]]
 

''Raw name(s)'' column 
The raw name list is automatically filled. In the case of this tutorial there is only one genome: '''NCBI36''' (Figure 10). 
''value:'' '''NCBI36'''
[[image:mg_hg18tohg19_raw_name.png|center|frame|Figure 10: Raw name list]]
Again, value is saved by closing the windows
 

'''''Import XML settings''''' 
In order to set the project with ease, user can import the settings using the XML file above. Please be careful about the VCF path, user must changes it directly on the xml file if he wants to use the import function.
 

'''''Conclusion''''' 
Finally, the screen should be like the one on Figure 11.
[[image:mg_hg18tohg19_vcf_loader.png|center|frame|Figure 11: VCF loader]]

===== Conclusion =====
The welcome screen should finally be similar to the Figure 12.
[[image:mg_hg18tohg19_welcome_screen.png|center|frame|Figure 12: Welcome screen]]

The "Create" button will create the project and will run the synchronization.

==== GRCh37/hg19 genes loading ====
To load a file, user has to do a right click on the left part of the track. Then to choose "Load Gene Track", a file chooser appears to select the file given in this tutorial. After having chosen the BED file, a new selection box appears (Figure 13).
[[image:mg_hg18tohg19_genome_selector_01.png|center|frame|Figure 13: Genome selection dialog for GRCh37/hg19 genes file]]

This box asks which genome is related to the BED file. Here, user has to choose "Feb 2009 (GFCh37/hg19)" option because the BED file contains information about that genome.
Gene file for GRCh37/hg19 reference has been loaded.

==== NCBI36/hg18 genes loading ====
The same operation as loading a gene files for GRCh37/hg19 reference genome. The only step changing is to choose the "Reference genome - hg18 (NCBI36)" option after the BED file selection (Figure 14)
[[image:mg_hg18tohg19_genome_selector_02.png|center|frame|Figure 14: Genome selection dialog for NCBI36/hg18 genes file]]

==== Conclusion ====
User can navigate into the different chromosomes and visualizes differences between both genomes using the stripes. All genes are perfectly synchronized and are display according to the meta-genome coordinates.

The Figure 15 shows an example of the result of this tutorial. It is possible to see deletions (in red) and insertions (in green) in the NCBI36/Hg18 reference genome compare to the GCh37/Hg19 reference genome. 
Chromosome: ''chr1'' 
Position: ''143,822,670'' 
[[image:mg_hg18tohg19_genplay_gene_example.png|center|thumb|800px|Figure 14: GenPlay-MG (chr1:143,822,670)]]

Multi-Genome Tutorial

2013-10-11T19:31:44Z

Nicolas: /* Getting started */

In order to set up and manage a Multi-Genome Project in Genplay, please refer to the following sections of the documentation:
* [[Documentation#Multi Genome Project|Loading a Multi Genome Project]]
* [[Documentation#Loading a Variant Layer|Loading a Variant Layer]]
* [[Documentation#Variant Layer Operations|Variant Layer operations]]

== Conversion between NCBI36/hg18 and GRCh37/hg19 ==
=== Description ===
This tutorial will explain how to display at the same time tracks mapped on genome assembly NCBI36/hg18 or GRCh37/hg19. In the example, user will be able to see all the modifications on the NCBI36/hg18 genome leading to the GRCh37/hg19 reference genome.

=== Files ===
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_settings.xml XML settings file]
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_sv.vcf.gz VCF file]
*[http://www.genplay.net/library/Human/Multi-Genome/hg18tohg19_tutorial_sv.vcf.gz.tbi Indexed VCF file (Tabix)]
*[http://www.genplay.net/library/Human/hg19/Gene_Annotation/RefSeq_From_UCSC_04-23-10(hg19).bed Refseq BED file for GRCh37/hg19]
*[http://www.genplay.net/library/Human/hg18/Gene_Annotation/RefSeq_From_UCSC_04-23-10(hg18).bed Refseq BED file for NCBI36/hg18]

=== Steps ===
==== Project settings ====
===== Project name =====
User must choose a name for a new project; here the name is ''GenPlay-MG – Reference genome tutorial'' (Figure 1).
[[image:mg_hg18tohg19_project_name.png|center|frame|Figure 1: Project name]]

===== Project assembly =====
According to bed files provided in this tutorial, the reference genome is GRCh37/hg19. User has to select the ''mammal'' clade, the ''human'' genome and the ''Feb 2009 (GRCh37/hg19)'' assembly as in Figure 2.
[[image:mg_hg18tohg19_project_assembly.png|center|frame|Figure 2: Project assembly]]

===== Chromosome selection =====
The VCF file is about Structural Variants and contains information for chromosomes 1 to 22 and chromosomes X and Y. User can select to load on or more chromosomes by clicking on the settings button next to the assembly name (Figure 3).
[[image:mg_hg18tohg19_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

===== VCF Loading =====
'''''Manually''''' 
To load VCF files , users must first fill the column lists and then select from the list the appropriate data. The VCF Loader appears after clicking on the ''Edit'' button from the welcome screen. The bottom left part of the VCF Loader contains the ''Column list edition'' section. User has to select a column and click on ''Edit'' button in order to show the associated list.
Only one VCF file is going to be loaded for this tutorial. The VCF file contains differences between the reference genome NCBI36/hg18 and the reference genome GRCh37/hg19.
 

''Group'' column 
This tutorial compares reference genome; a generic group name can be ''Reference genome''.
On the ''Group name list editor'', user clicks on the plus button to show the input text box and fills it (Figure 4). 
The ''Group name list editor'' should looks like the Figure 5 below. 
Once the values has been added to the list, it can be saved by closing the "Group name list editor window" 
''value:'' ''' Reference genome'''
<gallery widths=350px perrow=2>
image:mg_hg18tohg19_group_input.png|Figure 4: Group name input dialog
image:mg_hg18tohg19_group_editor.png|Figure 5: Group name editor
</gallery>
 

''Genome'' column 
The genome name is an Alias for the selected raw name. In this tutorial, the genome name is going to be '''Hg18'''.
On the ''Genome name list editor'', user clicks on the plus button to invoke the input text box and fills it (Figure 6). 
The ''Genome name list editor'' should looks like the Figure 7 below. 
Once the values has been added to the list, it can be saved by closing the "Genome name list editor window" 
''value:'' '''Hg18'''
<gallery widths=350px perrow=2>
image:mg_hg18tohg19_genome_input.png|Figure 6: Genome name input dialog
image:mg_hg18tohg19_genome_editor.png|Figure 7: Genome name editor
</gallery>
 

''Type'' column 
This field cannot be edited by the users. The provided VCF file is a Structural Variant type, user therefore has to choose '''SV''' (Figure 8). 
''value:'' '''SV'''
[[image:mg_hg18tohg19_type.png|center|frame|Figure 8: VCF type list]]
 

''File'' column 
Once the VCF file is downloaded, user has to open the ''File list editor'', user clicks on the plus button to show the file chooser dialog and choose the VCF file according to its location. 
''value:'' '''VCF path'''
[[image:mg_hg18tohg19_file_editor.png|center|frame|Figure 9: VCF File editor]]
 

''Raw name(s)'' column 
The raw name list is automatically filled. In the case of this tutorial there is only one genome: '''NCBI36''' (Figure 10). 
''value:'' '''NCBI36'''
[[image:mg_hg18tohg19_raw_name.png|center|frame|Figure 10: Raw name list]]
Again, value is saved by closing the windows
 

'''''Import XML settings''''' 
In order to set the project with ease, user can import the settings using the XML file above. Please be careful about the VCF path, user must changes it directly on the xml file if he wants to use the import function.
 

'''''Conclusion''''' 
Finally, the screen should be like the one on Figure 11.
[[image:mg_hg18tohg19_vcf_loader.png|center|frame|Figure 11: VCF loader]]

===== Conclusion =====
The welcome screen should finally be similar to the Figure 12.
[[image:mg_hg18tohg19_welcome_screen.png|center|frame|Figure 12: Welcome screen]]

The "Create" button will create the project and will run the synchronization.

==== GRCh37/hg19 genes loading ====
To load a file, user has to do a right click on the left part of the track. Then to choose "Load Gene Track", a file chooser appears to select the file given in this tutorial. After having chosen the BED file, a new selection box appears (Figure 13).
[[image:mg_hg18tohg19_genome_selector_01.png|center|frame|Figure 13: Genome selection dialog for GRCh37/hg19 genes file]]

This box asks which genome is related to the BED file. Here, user has to choose "Feb 2009 (GFCh37/hg19)" option because the BED file contains information about that genome.
Gene file for GRCh37/hg19 reference has been loaded.

==== NCBI36/hg18 genes loading ====
The same operation as loading a gene files for GRCh37/hg19 reference genome. The only step changing is to choose the "Reference genome - hg18 (NCBI36)" option after the BED file selection (Figure 14)
[[image:mg_hg18tohg19_genome_selector_02.png|center|frame|Figure 14: Genome selection dialog for NCBI36/hg18 genes file]]

==== Conclusion ====
User can navigate into the different chromosomes and visualizes differences between both genomes using the stripes. All genes are perfectly synchronized and are display according to the meta-genome coordinates.

The Figure 15 shows an example of the result of this tutorial. It is possible to see deletions (in red) and insertions (in green) in the NCBI36/Hg18 reference genome compare to the GCh37/Hg19 reference genome. 
Chromosome: ''chr1'' 
Position: ''143,822,670'' 
[[image:mg_hg18tohg19_genplay_gene_example.png|center|thumb|800px|Figure 14: GenPlay-MG (chr1:143,822,670)]]

Documentation

2013-10-11T19:23:46Z

Nicolas: /* Loading a Mask Layer */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 

=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#Filters|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>
==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

Documentation

2013-10-11T19:23:18Z

Nicolas: /* Loading a Variant Layer */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
[[image:Stripes.png|right|thumb|200px|CPG Islands Shown As Stripes On a Refseq Gene Layer]]
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 
=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
==== Add a Variant Layer ====
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|left|thumb|150px|Coordinate System chooser]]
The coordinate system of GenPlay can be changed by selecting one on the list located on the bottom right of the main frame. The default system is the one of the Meta Genome; the Reference Genome coordinate system is also available. The user can also choose the one of any of the loaded genome. This does not affect operation, only the red position numbers on the top of the frame as well as the position search bar on the bottom.
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|right|thumb|200px|Properties Dialog Button]]
In Multi-Genome Projects only, a new button appears on the bottom left of the frame. This button leads to the Multi-Genome Project Properties dialog allowing the user to visualize and handle the project settings. Right-clicking on the button opens a contextual menu offering shortcuts to the different sections of the properties dialog.
====== General ======
[[image:mg_general.png|right|thumb|300px|General Section]]
The General section is an overview of how the project has been loaded. Projects can be very complex, using many files and samples. This section reminds the user how the project has been set up.
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|Settings Section]]
The Settings section lets the user choose how to handle multi-genome various options.
* Properties Dialog
** Default section to open: the default section of the Multi-Genome Project Properties dialog to open when clicking the button.
* VCF Loader
** Default group text name: Default name for groups.
* Stripes transparency: Sets the transparency of stripes reprensenting variations.
* Global display settings
** Show legend: Allow to show the enabled variations and their colors into the track layer.
* Variant stripes settings
** Show filtered variation: Filtered variations can be shown but will be represented with a cross over their stripes.
** Show border of insertion: Insertion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show border of deletion: Deletion stripes have a specific border, it may help to recognize them easily when many layers are loaded, independantly of the color.
** Show nucleotides of insertion stripes: Added nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of deletion stripes: Deleted nucleotides will be retrieved from the VCF files if possible.
** Show nucleotides of SNP stripes: SNP nucleotides will be retrieved from the VCF files if possible.
* Reference stripes settings
** Show reference stripes: Stripes representing the reference genome can be either shown or hidden.
** Reference stripes color: Defines a color for reference stripes.
====== Files ======
The Files section lists all the VCF files loaded into GenPlay. Their information are separated into two categories:
* Information: the information part shows the name and the location of the file. It also segments the header of the VCF file for an easy reading and interpretation.
* Statistics: This part gives various descriptive statistics of the file and for each sample. All tables can be copied and pasted as regular text tab-delimited.
<gallery widths=350px height=150px perrow=2>
image:mg_file_info.png|File Information
image:mg_file_stat.png|File Statistics
</gallery>
====== Filters ======
The filters section is covered in the [[#Filters|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>
==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

Documentation

2013-10-10T21:18:14Z

Nicolas: /* Loading a Variant Layer */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
[[image:Stripes.png|right|thumb|200px|CPG Islands Shown As Stripes On a Refseq Gene Layer]]
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 
=== Loading a Variant Layer ===
==== Add a Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
==== Multi-Genome Features ====
Documentation in writing...
===== Select Coordinate System =====
[[image:mg_coordinate_chooser.png|right|thumb|150px|]]
===== Multi-Genome Project Properties =====
[[image:mg_option_button.png|center|thumb|200px|]]
====== General ======
[[image:mg_general.png|right|thumb|300px|]]
====== Settings ======
[[image:mg_settings.png|right|thumb|300px|]]
====== Files ======
[[image:mg_file_info.png|right|thumb|300px|]]
[[image:mg_file_stat.png|right|thumb|300px|]]
====== Filters ======
The filters section is covered in the [[#Filters|section below]].
 

=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>
==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

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Nicolas:

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Documentation

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Nicolas: /* Variant Layer Operations */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
[[image:Stripes.png|right|thumb|200px|CPG Islands Shown As Stripes On a Refseq Gene Layer]]
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 
=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
 
=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
==== Edit Variant Layer ====
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Edit Variant Layer Dialog]]
This feature will popup the same window used to load the Variant Layer offering the possibility to change the variation types to show.
==== Generate track statistics ====
This operations generates various statistics about loaded information. 
It also compares these statistics before and after applying any filters in order to see their effects.
==== Filters ====
Filters can be applied on Variant Layers, they interact directly on data found in the VCF in order to select on data of interest.
All filters are set in the Filters section of the Multi-Genome Project Properties dialog. 
Simply click on "Add" in order to create a new filter. As shown below, a new window appears to define the filter.
[[image:mg_filter_selection.png|center|thumb|800px|Filter selection dialog]]
* Layer(s): The layers affected by the filter.
* File: A filter is also file specific, if data to filter are separated over different files, several filters must be created.
* ID: A filter can be set on any ID defined on the header of the VCF. IDs can be of different types which affects the selection of the next steps.
* Genome(s): Any "FORMAT" ID will require to know which genome(s) is/are concerned by the filter.
* Operator: If more than one genome has been selected in the previous step, the operator will decide how the result from each genome will be processed in order to have a result for the whole line.
** And: The selected ID value from each genome must pass the filter.
** Or: At least one selected ID value must pass the filter.
** Sum: If the selected ID value is an integer, the sum value from each genome will be filtered.
** Mean: If the selected ID value is an integer, the mean value from all genomes will be filtered.
* Filter: This filter panel will change according to the selected ID type.
** String: The input value will be tested and the user has to choose if the value must be present or must not be present in the ID value.
** Number: The ID value is here tested using one of the given numeric operator against an input value. The ID value can also be tested against two input value using the secong part of the filter, the user then has to choose how both filters are handled.
** Flag: When the ID value is a flag, it reacts as boolean, meaning the value is here, or is not.
** Genotype: The genotype ID has a special filter editor in order to set it up more easily. The regular string editor can be found below. The genotype can be homozygote/heterofygote/phased/unphased.
<gallery widths=160px heights=160px perrow=4>
image:mg_filter_string.png|String Editor
image:mg_filter_number.png|Number Editor
image:mg_filter_flag.png|Flag Editor
image:mg_filter_genotype.png|Genotype Special Editor
</gallery>
==== Export as VCF ====
This operation exports all visible variations of the layer into a new VCF file. It includes filters meaning that it exports what can be seen on the layer.
==== Convert into variable window track ====
This operation converts the Variant Layer into a Microarray/Sequencing Layer. The new windows match the positions of the variation stripes. The score of the new windows can be set to any integer value present into the VCF lines. For haploid genomes, only one layer will be generated. For diploid genomes, the maternal and paternal alleles will be generated over two different layers.
==== Apply Genotype ====
Documentation in writing...

File:Mg filter selection.png

2013-10-10T20:57:42Z

Nicolas:

File:Mg filter genotype.png

2013-10-10T20:54:33Z

Nicolas:

File:Mg filter flag.png

2013-10-10T20:54:21Z

Nicolas:

File:Mg filter number.png

2013-10-10T20:54:09Z

Nicolas:

File:Mg filter string.png

2013-10-10T20:53:55Z

Nicolas:

Documentation

2013-10-08T19:30:27Z

Nicolas: /* Mask Layer Operations */

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
[[image:Stripes.png|right|thumb|200px|CPG Islands Shown As Stripes On a Refseq Gene Layer]]
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 
=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
 
=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
==== Apply Mask ====
Applying a mask means filtering the data that are not inside the windows of the mask. 
All information overlapping a mask window will be kept, everything else will be lost.
==== Invert Mask ====
This operation simply inverts all windows of the mask. All current windows become empty spaces, all empty spaces become windows.
 

=== Variant Layer Operations ===
Documentation in writing...

GenPlay File Formats

2013-10-07T15:12:07Z

Nicolas: /* Example */

This page presents the different file formats that can be loaded in GenPlay. Part of the information on this page is from the [http://genome.ucsc.edu/FAQ/ FAQ] of the UCSC genome browser. Don't hesitate to check it out for more details.

== .2bit format ==
=== Description ===
A .2bit file stores multiple DNA sequences (up to 4 Gb total) in a compact randomly-accessible format. The file contains masking information as well as the DNA itself.

The file begins with a 16-byte header containing the following fields:

# signature - the number 0x1A412743 in the architecture of the machine that created the file
# version - zero for now. Readers should abort if they see a version number higher than 0.
# sequenceCount - the number of sequences in the file.
# reserved - always zero for now

All fields are 32 bits unless noted. If the signature value is not as given, the reader program should byte-swap the signature and check if the swapped version matches. If so, all multiple-byte entities in the file will have to be byte-swapped. This enables these binary files to be used unchanged on different architectures.

The header is followed by a file index, which contains one entry for each sequence. Each index entry contains three fields:

# nameSize - a byte containing the length of the name field
# name - the sequence name itself, of variable length depending on nameSize
# offset - the 32-bit offset of the sequence data relative to the start of the file

The index is followed by the sequence records, which contain nine fields:

# dnaSize - number of bases of DNA in the sequence
# nBlockCount - the number of blocks of Ns in the file (representing unknown sequence)
# nBlockStarts - the starting position for each block of Ns
# nBlockSizes - the size of each block of Ns
# maskBlockCount - the number of masked (lower-case) blocks
# maskBlockStarts - the starting position for each masked block
# maskBlockSizes - the size of each masked block
# reserved - always zero for now
# packedDna - the DNA packed to two bits per base, represented as so: T - 00, C - 01, A - 10, G - 11.

The first base is in the most significant 2-bit byte; the last base is in the least significant 2 bits. For example, the sequence TCAG is represented as 00011011. The packedDna field is padded with 0 bits as necessary to take an even multiple of 32 bits in the file, which improves I/O performance on some machines.

=== Usage ===
The .2bit files can be loaded as sequence tracks.

To be loaded the file needs to have a '.2bit' extension.

== BAM / SAM format ==
=== Description ===
SAM (Sequence Alignment/Map) format is a generic format for storing large nucleotide sequence alignments.

BAM is a Binary version of the Sequence Alignment / Map (SAM) format.

More information about these file formats can be found [http://samtools.sourceforge.net/ here]

=== Usage ===
SAM files can be loaded as fixed window track.
BAM files can't be used directly in GenPlay but can be easily converted into SAM files using tools available on the Internet.

To be loaded a SAM file needs to either have a '.sam' extension or start with the following line:
track type=sam

== BED format ==
=== Description ===
BED format has three necessary fields and nine additional optional fields. The number of fields per line must be constant throughout any single set of data in an annotation track. The order of the optional fields requires that lower-numbered fields be filled if higher-numbered fields are used.

*The first three required BED fields are:
# chromosome - The name of the chromosome (e.g. chr1, chrX etc.) or scaffold (e.g. scaffold10461).
# Start - The beginning position of the feature in the chromosome or scaffold. The first base in a chromosome is numbered 0.
# End - The ending position of the feature in the chromosome or scaffold.
*The 9 additional optional BED fields are:
# name – This is the name of the gene.
# score - A score between 0 and 1000.
# strand - Defines the strand as either '+' (red) or '-' (blue).
# thickStart
# thickEnd
# itemRgb
# blockCount
# blockSizes
# blockStarts

=== Example ===
Here is an example of a bed file.

track type=bed geneDBURL="<nowiki>http://genome.ucsc.edu/cgi-bin/hgGene?org=Human&hgg_chrom=none&db=hg19&hgg_gene=</nowiki>"
chr1 11873 14409 uc001aaa.3 0 + 11873 11873 0 3 354,109,1189, 0,739,1347,
chr1 11873 14409 uc010nxq.1 0 + 12189 13639 0 3 354,127,1007, 0,721,1529,
chr1 11873 14409 uc010nxr.1 0 + 11873 11873 0 3 354,52,1189, 0,772,1347,
chr1 14362 16765 uc009vis.2 0 - 14362 14362 0 4 467,69,147,159, 0,607,1433,2244,

'''Note''':
The search URL "<nowiki>http://genome.ucsc.edu/cgi-bin/hgGene?org=Human&hgg_chrom=none&db=hg19&hgg_gene=</nowiki>" is included at the beginning of each BED file. It provides the URL that contains the description of the genes. One can form the complete URL by appending the name of the gene at the end of the search URL after the equal to sign. For example, "<nowiki>http://genome.ucsc.edu/cgi-bin/hgGene?org=Human&hgg_chrom=none&db=hg19&hgg_gene=uc001aaa.3</nowiki>" will provide information on WDR 78.

=== Usage ===
A Bed file with the first 3 required fields can generate stripes.
If the first four required fields are specified, the file can be loaded as a fixed window, a variable window or a repeat track.
If all the fields are specified the file can also be loaded as a gene track.

To be loaded a Bed file needs to either have a '.bed' extension or start with the following line:
track type=bed

== BedGraph format ==
=== Description ===
The BedGraph format is a really simple format useful to visualize windows on the genome. This windows can have a score.
The fields in a BedGraph file are the followings:
# Chromosome
# Window start position
# Window stop position
# Score

=== Example ===
track type =bedgraph
chr1 18598 19673 1
chr1 124987 125426 3
chr1 317653 318092 15
chr1 427014 428027 8

=== Usage ===
BedGraph files can be used to load fixed window and variable window track.

They can also be loaded as stripes

A valid BedGraph file needs to have a '.bgr' extension or should start with the following line:
track type=bedgraph

== Eland Extended format ==
=== Description ===
The Eland Extended files contain the result of read alignments on a reference genome.

Each line of an Eland Extended file contains the following fields:
# Sequence or read name
# Sequence
#
#* Either NM (No match found), QC (no matching done because of quality control failure (too many Ns)), RM (No matching done: repeat masked (may be seen if repeatFile.txt specified)
#* Or x:y:z where x, y, and z are the number of exact, single-error, and 2-error matches found
#
#* Either blank, if no matches found or if too many matches found
#* Or the following: BAC_plus_vector.fa:163022R1,170128F2,E_coli.fa:3909847R1 This says there are two matches to BAC_plus_vector.fa: one in the reverse direction starting at position 160322 with one error, one in the forward direction starting at position 170128 with two errors. There is also a single-error match to E_coli.fa.

=== Usage ===
The Eland Extended files can generate fixed window tracks.

The file needs to have a '.elx' extension or to start with the following line:
track type=eland_extended

== GenPlay Project format ==
=== Description ===
GenPlay Project files can contain an entire project from GenPlay. These files are extremely compact because they are saved as compressed binary files. This means that they can't be edited with a text editor.

=== Usage ===
An entire GenPlay project can be loaded or saved as a GenPlay Project file. The only tracks that are not included in the project files are the sequence tracks.

A project file must have a '.gen' extension.

== GdpGene format ==
=== Description ===
The GdpGene file format can be used to store information about genes. It is different from the BED format because it can store one score value for each exon.

The 7 mandatory fields of a GdpGene file are:

# name
# chromosome
# strand
# start
# stop
# exon starts (list of the exon start positions separated by commas and with no spaces or tabulations)
# exon stops (list of the exon stop positions separated by commas and with no spaces or tabulations)

There is also an optional field:
# exon scores (list of the exon scores separated by commas and with no spaces or tabulations)

=== Example ===
Here is an example of a GdpGene file:
track type=GdpGene name=Genes_Mouse_RefSeq_07-2007.txt
Xkr4 chr1 - 3204562 3661579 3204562,3411782,3660632 3207049,3411982,3661579
Rp1 chr1 - 4334223 4350473 4334223,4341990,4342282,4350280 4340172,4342162,4342918,4350473
Sox17 chr1 - 4481008 4486494 4481008,4483852,4485216,4486371 4482749,4483944,4486023,4486494

'''Note''': you can add a searchURL line to the GdpGene file. It works the same way as for a BED file, described [[#BED format|here]].

=== Usage ===
GdpGene files can be used to load gene tracks.

A valid GdpGene file needs either to have a '.gdp' extension or to have the following first line:
track type=GdpGene

== GFF format ==
=== Description ===
GFF stands for General Feature Format. GFF files must have 9 compulsory fields which are tab delimited. They are as follows:
# seqname - The name of the sequence. Must be a chromosome.
# source - The source code that was responsible for generation of this feature.
# feature - The name of this type of feature.
# start - The starting position of the feature in the sequence. The first base are 1 indexed.
# end - The ending position of the feature (inclusive).
# score - A score between 0 and 1000.
# strand - Valid entries include '+', '-', or '.' (for don't know/don't care).
# frame – Is a number between 0 and 2, if the feature is a coding exon and if the feature is not a coding exon, it is ‘.’.
# group - Lines belonging to the same group are linked together into a single item.
More information about this format can be found on [http://www.sanger.ac.uk/resources/software/gff Sanger website]

=== Usage ===
GFF files can be used to load repeats, variable and fixed window track as well as stripes.

A GFF file needs to have a '.gff' extension or to have the following first line:
##GFF

== GTF format ==
=== Description ===
GTF stands for Gene Transfer Format and is a stricter version of GFF. The first 8 GTF fields are the same as GFF. The group field has been expanded into a list of attributes. Each attribute is a type/value pair. Attributes must be terminated by a semi-colon. The gap between any two attributes must be exactly one space.
The attribute list must begin with the two required attributes:
# gene_id value - A GUID for the genomic source of the sequence
# transcript_id value - A GUID for the predicted transcript
Refer to the [http://www.sanger.ac.uk/resources/databases/encode/gencodeformat.html Sanger website] for further information.

=== Usage ===
Stripes, fixed and variable window, repeat and gene tracks can be generated from a GTF file.

A GTF files must either have a '.gtf' extension or to have the following first line:
##GTF

== Pair format ==
=== Description ===
This file format is used by NimbleGen.
The fields of a NimbleGen Pair file are:

# Image ID
# Gene Expression Option
# Sequence ID
# Probe ID
# Position
# X
# Y
# Match Index
# PM
# MM

Please refer to the documentation of [http://www.nimblegen.com/products/lit/NimbleScan_v2p6_UsersGuide.pdf NimbleScan] (page 114) for further information on this format.

=== Usage ===
The pair files can generate fixed window tracks.

The file needs to have a '.pair' extension to be loaded in GenPlay.

== PSL format ==
=== Description ===
PSL lines represent alignments, and are typically taken from files generated by BLAT or psLayout. All of the following fields are required on each data line within a PSL file:

# matches - Number of bases that match that aren’t repeats
# mismatches - Number of bases that don't match
# repeats - Number of bases that match but are part of repeats
# nbasecount - Number of 'N' bases
# numofinsertsquery - Number of inserts in query
# numofbaseinsertsquery - Number of bases inserted in query
# numofinsertstarget - Number of inserts in target
# numofbaseinsertsquery - Number of bases inserted in target
# strand - '+' or '-' for query strand. For translated alignments, second '+'or '-' is for genomic strand
# queryseqname - Query sequence name
# queryseqize - Query sequence size
# querystart - Alignment start position in query
# queryend - Alignment end position in query
# targetseqname - Target sequence name
# targetseqsize - Target sequence size
# targetstart - Alignment start position in target
# targetend - Alignment end position in target
# blockcount - Number of blocks in the alignment (a block contains no gaps)
# blocksizes - Comma-separated list of sizes of each block
# querystarts - Comma-separated list of starting positions of each block in query
# targetstarts - Comma-separated list of starting positions of each block in target

=== Usage ===
A PSL file can generate stripes. It can also be loaded as a fixed window, a variable window, a gene or a repeat track.

To be loaded a PSL file needs either to have a '.psl' extension or to have the following first line:
track type=psl

== SOAPsnp format ==
=== Description ===
The SOAPsnp files contain the result of a resequencing utility that can assemble consensus sequence for the genome of a newly sequenced individual based on the alignment of raw sequencing reads on a known reference. The SNPs can then be identified on the consensus sequence through the comparison with the reference. More information about this file format can be found [http://soap.genomics.org.cn/soapsnp.html here]

The result of SOAPsnp has 17 columns:

# Chromosome ID
# Coordinate on chromosome, start from 1
# Reference genotype
# Consensus genotype
# Quality score of consensus genotype
# Best base
# Average quality score of best base
# Count of uniquely mapped best base
# Count of all mapped best base
# Second best bases
# Average quality score of second best base
# Count of uniquely mapped second best base
# Count of all mapped second best base
# Sequencing depth of the site
# Rank sum test p_value
# Average copy number of nearby region
# Whether the site is a dbSNP.

=== Usage ===
The SOAPsnp files can be used to generate SNPs tracks.

Be sure that the file has a '.soapsnp' extension or starts with the following line:
track type=SOAPsnp

== WIG format ==
=== Description ===
Wiggle format (WIG) allows the display of continuous-valued data in a track format.
Please refer to this [http://genome.ucsc.edu/goldenPath/help/wiggle.html page] for the whole description of a wiggle file.

==== General Structure ====
Wiggle format is line-oriented. For wiggle custom tracks, the first line must be a track definition line, which designates the track as a wiggle track and adds a number of options for controlling the default display.

Wiggle format is composed of declaration lines and data lines. There are two options for formatting wiggle data: '''variableStep''' and '''fixedStep'''. These formats were developed to allow the file to be written as compactly as possible.

* '''variableStep''' is for data with irregular intervals between new data points and is the more commonly used wiggle format. It begins with a declaration line and is followed by two columns containing chromosome positions and data values:

variableStep chrom=chrN [span=windowSize]
chromStartA dataValueA
chromStartB dataValueB
... etc ... ... etc ...

The declaration line starts with the word variableStep and is followed by a specification for a chromosome. The optional span parameter (default: span=1) allows data composed of contiguous runs of bases with the same data value to be specified more succinctly. The span begins at each chromosome position specified and indicates the number of bases that data value should cover.

For example, this variableStep specification:

variableStep chrom=chr2
300701 12.5
300702 12.5
300703 12.5
300704 12.5
300705 12.5

is equivalent to:

variableStep chrom=chr2 span=5
300701 12.5

Both versions display a value of 12.5 at position 300701-300705 on chromosome 2.

* '''fixedStep''' is for data with regular intervals between new data values and is the more compact wiggle format. It begins with a declaration line and is followed by a single column of data values:

fixedStep chrom=chrN start=position step=stepInterval [span=windowSize]
dataValue1
dataValue2
... etc ...

The declaration line starts with the word fixedStep and includes specifications for chromosome, start coordinate, and step size. The span specification has the same meaning as in variableStep format.

For example, this fixedStep specification:

fixedStep chrom=chr3 start=400601 step=100
11
22
33

displays the values 11, 22, and 33 as single-base regions on chromosome 3 at positions 400601, 400701, and 400801, respectively.

Adding span=5 to the declaration line:

fixedStep chrom=chr3 start=400601 step=100 span=5
11
22
33

causes the values 11, 22, and 33 to be displayed as 5-base regions on chromosome 3 at positions 400601-400605, 400701-400705, and 400801-400805, respectively.

Note that for both '''variableStep''' and '''fixedStep''' formats, the same span must be used throughout the dataset. If no span is specified, the default span of one is used.

=== Usage ===
A wiggle file can generate fixed or variable window tracks as well as stripes.

Be sure that the file extension is '.wig' or that the file starts with the following line:
track type=wiggle

GenPlay File Formats

2013-10-07T13:33:52Z

Nicolas: /* Example */

This page presents the different file formats that can be loaded in GenPlay. Part of the information on this page is from the [http://genome.ucsc.edu/FAQ/ FAQ] of the UCSC genome browser. Don't hesitate to check it out for more details.

== .2bit format ==
=== Description ===
A .2bit file stores multiple DNA sequences (up to 4 Gb total) in a compact randomly-accessible format. The file contains masking information as well as the DNA itself.

The file begins with a 16-byte header containing the following fields:

# signature - the number 0x1A412743 in the architecture of the machine that created the file
# version - zero for now. Readers should abort if they see a version number higher than 0.
# sequenceCount - the number of sequences in the file.
# reserved - always zero for now

All fields are 32 bits unless noted. If the signature value is not as given, the reader program should byte-swap the signature and check if the swapped version matches. If so, all multiple-byte entities in the file will have to be byte-swapped. This enables these binary files to be used unchanged on different architectures.

The header is followed by a file index, which contains one entry for each sequence. Each index entry contains three fields:

# nameSize - a byte containing the length of the name field
# name - the sequence name itself, of variable length depending on nameSize
# offset - the 32-bit offset of the sequence data relative to the start of the file

The index is followed by the sequence records, which contain nine fields:

# dnaSize - number of bases of DNA in the sequence
# nBlockCount - the number of blocks of Ns in the file (representing unknown sequence)
# nBlockStarts - the starting position for each block of Ns
# nBlockSizes - the size of each block of Ns
# maskBlockCount - the number of masked (lower-case) blocks
# maskBlockStarts - the starting position for each masked block
# maskBlockSizes - the size of each masked block
# reserved - always zero for now
# packedDna - the DNA packed to two bits per base, represented as so: T - 00, C - 01, A - 10, G - 11.

The first base is in the most significant 2-bit byte; the last base is in the least significant 2 bits. For example, the sequence TCAG is represented as 00011011. The packedDna field is padded with 0 bits as necessary to take an even multiple of 32 bits in the file, which improves I/O performance on some machines.

=== Usage ===
The .2bit files can be loaded as sequence tracks.

To be loaded the file needs to have a '.2bit' extension.

== BAM / SAM format ==
=== Description ===
SAM (Sequence Alignment/Map) format is a generic format for storing large nucleotide sequence alignments.

BAM is a Binary version of the Sequence Alignment / Map (SAM) format.

More information about these file formats can be found [http://samtools.sourceforge.net/ here]

=== Usage ===
SAM files can be loaded as fixed window track.
BAM files can't be used directly in GenPlay but can be easily converted into SAM files using tools available on the Internet.

To be loaded a SAM file needs to either have a '.sam' extension or start with the following line:
track type=sam

== BED format ==
=== Description ===
BED format has three necessary fields and nine additional optional fields. The number of fields per line must be constant throughout any single set of data in an annotation track. The order of the optional fields requires that lower-numbered fields be filled if higher-numbered fields are used.

*The first three required BED fields are:
# chromosome - The name of the chromosome (e.g. chr1, chrX etc.) or scaffold (e.g. scaffold10461).
# Start - The beginning position of the feature in the chromosome or scaffold. The first base in a chromosome is numbered 0.
# End - The ending position of the feature in the chromosome or scaffold.
*The 9 additional optional BED fields are:
# name – This is the name of the gene.
# score - A score between 0 and 1000.
# strand - Defines the strand as either '+' (red) or '-' (blue).
# thickStart
# thickEnd
# itemRgb
# blockCount
# blockSizes
# blockStarts

=== Example ===
Here is an example of a bed file.

track type=bed
geneDBURL="<nowiki>http://genome.ucsc.edu/cgi-bin/hgGene?org=Human&hgg_chrom=none&db=hg19&hgg_gene=</nowiki>"
chr1 11873 14409 uc001aaa.3 0 + 11873 11873 0 3 354,109,1189, 0,739,1347,
chr1 11873 14409 uc010nxq.1 0 + 12189 13639 0 3 354,127,1007, 0,721,1529,
chr1 11873 14409 uc010nxr.1 0 + 11873 11873 0 3 354,52,1189, 0,772,1347,
chr1 14362 16765 uc009vis.2 0 - 14362 14362 0 4 467,69,147,159, 0,607,1433,2244,

'''Note''':
The search URL "<nowiki>http://genome.ucsc.edu/cgi-bin/hgGene?org=Human&hgg_chrom=none&db=hg19&hgg_gene=</nowiki>" is included at the beginning of each BED file. It provides the URL that contains the description of the genes. One can form the complete URL by appending the name of the gene at the end of the search URL after the equal to sign. For example, "<nowiki>http://genome.ucsc.edu/cgi-bin/hgGene?org=Human&hgg_chrom=none&db=hg19&hgg_gene=uc001aaa.3</nowiki>" will provide information on WDR 78.

=== Usage ===
A Bed file with the first 3 required fields can generate stripes.
If the first four required fields are specified, the file can be loaded as a fixed window, a variable window or a repeat track.
If all the fields are specified the file can also be loaded as a gene track.

To be loaded a Bed file needs to either have a '.bed' extension or start with the following line:
track type=bed

== BedGraph format ==
=== Description ===
The BedGraph format is a really simple format useful to visualize windows on the genome. This windows can have a score.
The fields in a BedGraph file are the followings:
# Chromosome
# Window start position
# Window stop position
# Score

=== Example ===
track type =bedgraph
chr1 18598 19673 1
chr1 124987 125426 3
chr1 317653 318092 15
chr1 427014 428027 8

=== Usage ===
BedGraph files can be used to load fixed window and variable window track.

They can also be loaded as stripes

A valid BedGraph file needs to have a '.bgr' extension or should start with the following line:
track type=bedgraph

== Eland Extended format ==
=== Description ===
The Eland Extended files contain the result of read alignments on a reference genome.

Each line of an Eland Extended file contains the following fields:
# Sequence or read name
# Sequence
#
#* Either NM (No match found), QC (no matching done because of quality control failure (too many Ns)), RM (No matching done: repeat masked (may be seen if repeatFile.txt specified)
#* Or x:y:z where x, y, and z are the number of exact, single-error, and 2-error matches found
#
#* Either blank, if no matches found or if too many matches found
#* Or the following: BAC_plus_vector.fa:163022R1,170128F2,E_coli.fa:3909847R1 This says there are two matches to BAC_plus_vector.fa: one in the reverse direction starting at position 160322 with one error, one in the forward direction starting at position 170128 with two errors. There is also a single-error match to E_coli.fa.

=== Usage ===
The Eland Extended files can generate fixed window tracks.

The file needs to have a '.elx' extension or to start with the following line:
track type=eland_extended

== GenPlay Project format ==
=== Description ===
GenPlay Project files can contain an entire project from GenPlay. These files are extremely compact because they are saved as compressed binary files. This means that they can't be edited with a text editor.

=== Usage ===
An entire GenPlay project can be loaded or saved as a GenPlay Project file. The only tracks that are not included in the project files are the sequence tracks.

A project file must have a '.gen' extension.

== GdpGene format ==
=== Description ===
The GdpGene file format can be used to store information about genes. It is different from the BED format because it can store one score value for each exon.

The 7 mandatory fields of a GdpGene file are:

# name
# chromosome
# strand
# start
# stop
# exon starts (list of the exon start positions separated by commas and with no spaces or tabulations)
# exon stops (list of the exon stop positions separated by commas and with no spaces or tabulations)

There is also an optional field:
# exon scores (list of the exon scores separated by commas and with no spaces or tabulations)

=== Example ===
Here is an example of a GdpGene file:
track type=GdpGene name=Genes_Mouse_RefSeq_07-2007.txt
Xkr4 chr1 - 3204562 3661579 3204562,3411782,3660632 3207049,3411982,3661579
Rp1 chr1 - 4334223 4350473 4334223,4341990,4342282,4350280 4340172,4342162,4342918,4350473
Sox17 chr1 - 4481008 4486494 4481008,4483852,4485216,4486371 4482749,4483944,4486023,4486494

'''Note''': you can add a searchURL line to the GdpGene file. It works the same way as for a BED file, described [[#BED format|here]].

=== Usage ===
GdpGene files can be used to load gene tracks.

A valid GdpGene file needs either to have a '.gdp' extension or to have the following first line:
track type=GdpGene

== GFF format ==
=== Description ===
GFF stands for General Feature Format. GFF files must have 9 compulsory fields which are tab delimited. They are as follows:
# seqname - The name of the sequence. Must be a chromosome.
# source - The source code that was responsible for generation of this feature.
# feature - The name of this type of feature.
# start - The starting position of the feature in the sequence. The first base are 1 indexed.
# end - The ending position of the feature (inclusive).
# score - A score between 0 and 1000.
# strand - Valid entries include '+', '-', or '.' (for don't know/don't care).
# frame – Is a number between 0 and 2, if the feature is a coding exon and if the feature is not a coding exon, it is ‘.’.
# group - Lines belonging to the same group are linked together into a single item.
More information about this format can be found on [http://www.sanger.ac.uk/resources/software/gff Sanger website]

=== Usage ===
GFF files can be used to load repeats, variable and fixed window track as well as stripes.

A GFF file needs to have a '.gff' extension or to have the following first line:
##GFF

== GTF format ==
=== Description ===
GTF stands for Gene Transfer Format and is a stricter version of GFF. The first 8 GTF fields are the same as GFF. The group field has been expanded into a list of attributes. Each attribute is a type/value pair. Attributes must be terminated by a semi-colon. The gap between any two attributes must be exactly one space.
The attribute list must begin with the two required attributes:
# gene_id value - A GUID for the genomic source of the sequence
# transcript_id value - A GUID for the predicted transcript
Refer to the [http://www.sanger.ac.uk/resources/databases/encode/gencodeformat.html Sanger website] for further information.

=== Usage ===
Stripes, fixed and variable window, repeat and gene tracks can be generated from a GTF file.

A GTF files must either have a '.gtf' extension or to have the following first line:
##GTF

== Pair format ==
=== Description ===
This file format is used by NimbleGen.
The fields of a NimbleGen Pair file are:

# Image ID
# Gene Expression Option
# Sequence ID
# Probe ID
# Position
# X
# Y
# Match Index
# PM
# MM

Please refer to the documentation of [http://www.nimblegen.com/products/lit/NimbleScan_v2p6_UsersGuide.pdf NimbleScan] (page 114) for further information on this format.

=== Usage ===
The pair files can generate fixed window tracks.

The file needs to have a '.pair' extension to be loaded in GenPlay.

== PSL format ==
=== Description ===
PSL lines represent alignments, and are typically taken from files generated by BLAT or psLayout. All of the following fields are required on each data line within a PSL file:

# matches - Number of bases that match that aren’t repeats
# mismatches - Number of bases that don't match
# repeats - Number of bases that match but are part of repeats
# nbasecount - Number of 'N' bases
# numofinsertsquery - Number of inserts in query
# numofbaseinsertsquery - Number of bases inserted in query
# numofinsertstarget - Number of inserts in target
# numofbaseinsertsquery - Number of bases inserted in target
# strand - '+' or '-' for query strand. For translated alignments, second '+'or '-' is for genomic strand
# queryseqname - Query sequence name
# queryseqize - Query sequence size
# querystart - Alignment start position in query
# queryend - Alignment end position in query
# targetseqname - Target sequence name
# targetseqsize - Target sequence size
# targetstart - Alignment start position in target
# targetend - Alignment end position in target
# blockcount - Number of blocks in the alignment (a block contains no gaps)
# blocksizes - Comma-separated list of sizes of each block
# querystarts - Comma-separated list of starting positions of each block in query
# targetstarts - Comma-separated list of starting positions of each block in target

=== Usage ===
A PSL file can generate stripes. It can also be loaded as a fixed window, a variable window, a gene or a repeat track.

To be loaded a PSL file needs either to have a '.psl' extension or to have the following first line:
track type=psl

== SOAPsnp format ==
=== Description ===
The SOAPsnp files contain the result of a resequencing utility that can assemble consensus sequence for the genome of a newly sequenced individual based on the alignment of raw sequencing reads on a known reference. The SNPs can then be identified on the consensus sequence through the comparison with the reference. More information about this file format can be found [http://soap.genomics.org.cn/soapsnp.html here]

The result of SOAPsnp has 17 columns:

# Chromosome ID
# Coordinate on chromosome, start from 1
# Reference genotype
# Consensus genotype
# Quality score of consensus genotype
# Best base
# Average quality score of best base
# Count of uniquely mapped best base
# Count of all mapped best base
# Second best bases
# Average quality score of second best base
# Count of uniquely mapped second best base
# Count of all mapped second best base
# Sequencing depth of the site
# Rank sum test p_value
# Average copy number of nearby region
# Whether the site is a dbSNP.

=== Usage ===
The SOAPsnp files can be used to generate SNPs tracks.

Be sure that the file has a '.soapsnp' extension or starts with the following line:
track type=SOAPsnp

== WIG format ==
=== Description ===
Wiggle format (WIG) allows the display of continuous-valued data in a track format.
Please refer to this [http://genome.ucsc.edu/goldenPath/help/wiggle.html page] for the whole description of a wiggle file.

==== General Structure ====
Wiggle format is line-oriented. For wiggle custom tracks, the first line must be a track definition line, which designates the track as a wiggle track and adds a number of options for controlling the default display.

Wiggle format is composed of declaration lines and data lines. There are two options for formatting wiggle data: '''variableStep''' and '''fixedStep'''. These formats were developed to allow the file to be written as compactly as possible.

* '''variableStep''' is for data with irregular intervals between new data points and is the more commonly used wiggle format. It begins with a declaration line and is followed by two columns containing chromosome positions and data values:

variableStep chrom=chrN [span=windowSize]
chromStartA dataValueA
chromStartB dataValueB
... etc ... ... etc ...

The declaration line starts with the word variableStep and is followed by a specification for a chromosome. The optional span parameter (default: span=1) allows data composed of contiguous runs of bases with the same data value to be specified more succinctly. The span begins at each chromosome position specified and indicates the number of bases that data value should cover.

For example, this variableStep specification:

variableStep chrom=chr2
300701 12.5
300702 12.5
300703 12.5
300704 12.5
300705 12.5

is equivalent to:

variableStep chrom=chr2 span=5
300701 12.5

Both versions display a value of 12.5 at position 300701-300705 on chromosome 2.

* '''fixedStep''' is for data with regular intervals between new data values and is the more compact wiggle format. It begins with a declaration line and is followed by a single column of data values:

fixedStep chrom=chrN start=position step=stepInterval [span=windowSize]
dataValue1
dataValue2
... etc ...

The declaration line starts with the word fixedStep and includes specifications for chromosome, start coordinate, and step size. The span specification has the same meaning as in variableStep format.

For example, this fixedStep specification:

fixedStep chrom=chr3 start=400601 step=100
11
22
33

displays the values 11, 22, and 33 as single-base regions on chromosome 3 at positions 400601, 400701, and 400801, respectively.

Adding span=5 to the declaration line:

fixedStep chrom=chr3 start=400601 step=100 span=5
11
22
33

causes the values 11, 22, and 33 to be displayed as 5-base regions on chromosome 3 at positions 400601-400605, 400701-400705, and 400801-400805, respectively.

Note that for both '''variableStep''' and '''fixedStep''' formats, the same span must be used throughout the dataset. If no span is specified, the default span of one is used.

=== Usage ===
A wiggle file can generate fixed or variable window tracks as well as stripes.

Be sure that the file extension is '.wig' or that the file starts with the following line:
track type=wiggle

Documentation

2013-10-04T21:10:02Z

Nicolas:

== Getting started ==
=== Starting GenPlay ===
GenPlay is freely available at http://www.genplay.net/wiki/index.php/Web_Start To start the software, click the button corresponding to the amount of memory that you wish to allocate to the Java virtual machine.

The amount of memory determines how many layers you will be able to load simultaneously. The programming philosophy behind GenPlay is to provide fast performances once the data is loaded. To achieve that goal the entire genome need to be loaded in memory for multiple layers at the same time. This results in high quality performance, but requires a lot of memory. The amount of memory needed per layer depends on the genome, the layer type, the window size, the data precision etc.

You should generally choose as much memory as you can afford on your system (generally about 70% of the total RAM memory that exists on your system). For mammalian genomes we recommend allocating at least 4 GB of RAM although you should be able to load a couple of genome-wide layers with 1GB or 1.5GB of RAM. Selecting analysis of only one chromosome at a time will drastically reduce the memory requirement and should allow you to load many layers at very high resolutions. Layers loaded in GenPlay can also be compressed as explained later in this documentation.

The amount of RAM memory available to GenPlay is displayed in the lower right corner of the screen.
 
=== The Welcome screen ===
The welcome screen is the first screen of GenPlay-MG and allow user to create or to load a project.

==== New Project ====
In order to create a new project, users must give it a name as shown in Figure 1.
[[image:mg_basics_project name.png|center|frame|Figure 1: Text field to define the project name]]

The second step is to choose a reference genome. Users can choose it using the different list according to the clade, the genome and the assembly (Figure 2).
[[image:mg_basics_assembly_chooser.png|center|frame|Figure 2: Assembly chooser]]

Several chromosomes are available for each assembly but users can choose to select only some of them. 
To open the chromosome chooser (Figure 3), users have to click on the tools button next to the assembly name.
[[image:mg_basics_chromosome_chooser.png|center|frame|Figure 3: Chromosome chooser]]

The third and last step is to choose between a ''Simple Genome Project'' and a ''Multi Genome Project''. If the multi genome project option is selected, the welcome screen should be as the one shown in Figure 4.
[[image:mg_basics_empty_welcome_screen.png|center|frame|Figure 4: Empty welcome screen for multi-genome project]]
===== Single Genome Project =====

===== Multi Genome Project =====
====== Introduction ======
====== VCF Files ======
VCF files describe differences between genomes. Usually, it concerns differences between one or several genomes of interest and the reference genome used for the mapping process. VCF files define multiple type of variations; GenPlay is able to read and represent the followings:
* InDels
* SNPs
* SV (Structural Variation)

A complete description of VCF files is given on the 1000 genomes project website: 
[http://www.1000genomes.org/wiki/analysis/variant-call-format/vcf-variant-call-format-version-42 Variant Call Format specification] 

====== Tabix ======
: 1. Introduction
VCF files contain a lot of information which makes the scanning (loading) processes longer. 
In order to increase the scanning efficiency, VCF files have to be compressed and indexed. The compression is done using BGZip and the indexing with Tabix. 
[http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages] 
[http://sourceforge.net/projects/samtools/files/tabix/ Tabix download]

: 2. VCF files indexing methods
:: 2.1. Using GenPlay
GenPlay is now able to compress and index VCF files using the VCF Loader. 
The way the VCF Loader works is explained below. When you want to select the compressed file (.vcf.gz), simply select the VCF file (.vcf) instead. You may need to change the file extension filter in the file chooser in order to see .vcf files. 
GenPlay will look then for compressed/indexed files at the same location, if nothing is found, it will offer to compress and index the selected VCF file (Figure 1).
[[image:mg_vcf_loader_compress_index.png|center|frame|Figure 1: VCF Loader compress/index]]

It is fully automatic and non-platform dependent (works on Windows, Linux and Mac).

:: 2.2. Manually
First, please note the following process must be performed in either Linux or Mac environments. 
Each VCF files must be first compress to a BGZF (.bgz file) format. Tabix provides a tool to perform the compression.
After compression, VCF files must be indexed using the associated command.
Once Tabix is installed, two commands are necessary to perform the indexation.
 

Available commands from the Tabix folder: 
''bgzip -f VCF_PATH;'' 
''tabix –p vcf VCF_PATH;''
 
For example, a VCF file named my_vcf.vcf located in the same folder as Tabix can be indexed with the following commands (Figure 2): 
''bgzip -f ./my_vcf.vcf;'' 
''tabix –p vcf ./my_vcf.vcf.gz;''
[[image:mg_basics_indexation_commands.png|center|frame|Figure 2: VCF file indexation command]]
 
'''Note:''' the first command '''replaces''' the current VCF file by the compressed VCF file (.vcf.gz). The second command '''creates''' the indexed VCF file in the current folder (.vcf.gz.tbi). 
More options are available on [http://samtools.sourceforge.net/tabix.shtml Tabix manual reference pages].

====== The VCF Loader ======
: 1. Introduction
The VCF Loader is the most important part of multi-genome project settings. It allows users to load all necessary VCF files and to define how to extract information from them. It appears when users click on the "Edit" button from the welcome screen. 
The Figure 3 shows an empty VCF Loader screen.
[[image:Mg_welcome_screen_vcf_loader.png|center|frame|Figure 3: VCF loader]]

GenPlay-MG does not use directly the VCF file, it uses a compress version of it (.gz). Moreover, GenPlay-MG also needs the compress VCF file to be indexed with Tabix. Both file versions must be in the '''same folder''' and must have the '''same name''', only file extensions differ (.gz and .tbi).
In order to use GenPlay to generate additional files, please refer to the [[#Tabix|section above]].

The user can add or remove rows by right clicking on the table.

: 2. Columns description
'''''File''''' 
This column refers to the VCF file path. Once loaded, the raw name column is automatically filled with every raw genome name contained in the selected VCF file. 
'''''Raw name''''' 
The ''Raw name'' column list is automatically filled when a VCF file has been chosen. That list contains every genotype headers contained inside the selected VCF file. Because Genome names might be difficult to remembers, GenPlay-MG offers users the option of adding another name (an alias) using the ''Genome'' column. 
'''''Nickname''''' 
The ''Nickname'' column allows users to associate an alias to the selected genome. This alias will appear in GenPlay-MG and can be useful because genome names in VCF files are often non descriptive numbers that can be hard to remember. 
'''''Group''''' 
Users can gather genomes by group. Group names are used to distinguish genomes and to perform some specific functionalities. 

: 3. Columns edition 
''Group'', ''Nickname'' and ''File'' column have their own editable list.To edit a cell, click on it, go over the item you want to edit and choose one of the following action: 
- Add (green symbol on empty item) 
- Edit (pen symbol on an item) 
- Delete (red symbol on an item) 

That way, users can set up all columns before starting (or at the same time) to fill the table. 

'''Note: ''' The ''Raw name(s)'' column is automatically filled with genome name from the selected VCF file, that column cannot be edited manually.

====== Import/Export ======
Once a project has been set up, it can be saved using the import/export function. Pressing the export button saves an XML files to the hard drive. This XML file can then be imported to reload the project.

The XML file structure is simple. Each row are stored in ''row'' mark containing every attribute names such as ''group'', ''genome, ''file'' and ''raw_name''. The settings file is formatted as shown in Figure 4.
[[image:mg_basics_xml_settings.png|center|frame|Figure 4: XML file settings]]

'''Note:''' If the user moves the VCF files or changes one of its genotype headers, the XML file will not work anymore. User has to modify ''file'' and/or ''raw_name'' attribute values. 

==== Load Project ====
Documentation in writing...
 
== GUI Overview ==
[[image:gui_overview.png|right|thumb|200px|GUI Overview 1.Ruler 2.Track List 3.Control Panel 4.Status Bar]]

GenPlay main window is divided in 4 main parts:
# Ruler
# Track List
# Control Panel
# Status Bar
 
=== Ruler ===
The ruler shows the coordinates of the current displayed position.

[[image:ruler.png|left|thumb|500px|Ruler 1.Option Button 2.Absolute Positions 3.Relative Positions]]

==== General Option Button ====
The button on the left of the ruler opens the pop-up menu with all the general options.

==== Absolute Positions ====
The numbers written in red on top of the ruler are the absolute position on the selected chromosome or scaffold.

The number on the left is the position of the first displayed base. This value can be negative.

The number in the middle is the position of the red line. This value can go from 0 to the length of the current chromosome or scaffold as specified in the chromosome configuration file.

The value on the right is the last displayed position. This value range from 1 to 2*(chromosome length).

==== Relative Position ====
The numbers written in black on the second line represent the distance from the middle in base pair.
 
=== Track List ===
The track list is the cornerstone of the GUI. From here you can load layers and execute operations.

The tracks are divided into two parts.

On the left, there is the track handler that becomes highlighted when the mouse is over it. By right clicking on the track handler, a contextual menu appears with all the operations that can be executed on the track and its layer(s).

On the right, the data can be visualized.
 
=== Control Panel ===
[[image:control_panel.png|right|thumb|500px|Control Panel 1.Position Bar 2.Zoom Bar 3.Chromosome Box 4.Position Text Field]]
The control panel is divided into 4 parts:
# Position Bar: the position bar allows you to change the position of the current displayed windows
# Zoom Bar: use the zoom bar to modify the level of zoom
# Chromosome Box: set the selected chromosome with the chromosome box
# Position Text Field: the position text field follows the format of the UCSC Genome Browser position field so it is easy to copy and paste the position from one browser to the other
 
=== Status Bar ===
[[image:status_bar.png|right|thumb|500px|Status Bar 1.Progress Bar 2.Stop Button 3.Operation Description 4.Memory Bar]]
The status bar helps monitor the progress of the current operation as well as memory usage. It is divided into 4 sub-components:
# Progress bar, shows the level of completion of the current operation
# Stop button, allows users to stop the current operation. If the button is not bright red the operation can't be stopped
# Operation description, displays a short text describing the current operation as well as the elapsed time from the beginning of the operation
# Memory bar, shows the amount of memory used and the amount of memory available. Make sure that you have enough memory before starting a new operation. You can delete or compress layers to free up memory.
 
== Browsing the Genome ==
=== Changing the Position ===
You can change the position of the displayed window by:
# Dragging any track on the left or on the right with the left button of the mouse
# Clicking with the middle button of the mouse inside a track and then moving the cursor on the left or on the right of the middle red line
# Moving the knob of the position bar on the control panel
# Changing the value of the position text field on the control panel
# Using the keyboard left and right arrows
# Double-clicking on a track where you want to center the view
 
=== Changing the Chromosomes ===
You can switch the selected chromosome by:
# Changing the selection in the chromosome box on the control panel
# Changing the text of the position text field on the control panel
 
=== Changing the Zoom ===
The level of the zoom can be modified by:
# Wheeling up or down inside a track with the mouse wheel
# Using the zoom bar on the control panel
# Changing the text of the position text field on the control panel
 
== Loading a Layer ==
=== Introduction ===
The layers are the way to show information from files. They can represent information in different manners. 
A layer is created from a track, each track can contain one or several layers. 
To load a layer in a track, right click on its handler (the blue part on the left of the track). This opens a contextual menu with the different actions available on the track.
The menu of a track empty of layer looks like the one in figure 1. 
By clicking "Add Layer" appears a dialog to select one of the different layer type GenPlay offers (Figure 2). 
Examples of layers that can be loaded in GenPlay are available for download from the GenPlay Library accessible from the GenPlay.net website.

<gallery widths=350px perrow=2>
image:add_layer.png|Figure 1: Track Contextual Menu
image:layer_type.png|Figure 2: Layer Types
</gallery>
 
=== Loading a Sequencing/Microarray Layer ===
The Sequencing/Microarray layer allows the visualization of windows of variable/fix sizes with a score associated to these windows.
Select the “Sequencing/Microarray Layer” option. This opens up a file chooser dialog box. Load the file of your choice from the list of available window files and click the open button.

Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequencing/microarray layer.

This opens a new dialog to set different parameters for the new layer (as shown on the figure below). The dialog is separated in 6 sections detailed below.
[[image:Add_layer_seq_micro.png|right|thumb|300px|New Layer Settings Dialog]]

==== Layer Name ====
Gives a name to the layer.

==== Bin ====
By default, the windows generated in sequencing/microarray layer have a variable size. It represents very precisely the content of the file. 
For some other purposes, users may want to have fixed windows size. They are useful to represent the results of many types of experiments including, but not limited to: CHIP-seq, RNA seq, and TimEX-seq. Files containing the results of alignments (SAM, bowtie, Eland) and files containing already created bin lists (bed, bgr, etc.) can be loaded using this option. In the case of alignment files, bin lists will be created on the fly as described below. Files containing the results of micro-array experiments can also be loaded as long as they are in one of the accepted formats.
It lowers the resolution but usually offers better memory usage. 
This is implemented here by enabling the "Bin Data" option. The "Bin Size" field will then be available in order to give the size of the windows in base pairs. 

'''Important Note:''' A bin size of 1 bp will use a lot of memory. According to the experiment, it may be more efficient to disable the bin data option and stay in variable window size mode.

==== Score Calculation ====
[[image:score_calculation_methods.png|right|thumb|100px|Name and Score Calculation]]
It can happen that files contain overlapping windows. In this case, GenPlay splits them into smaller windows using a simple algorithm. 
This algorithm can be chosen in that section offering the following possibilities: 
* Addition
* Average
* Maximum
* Minimum

Some examples are shown in the sections below for both [[#For non bined layer|non bined]] and [[#For bined layer|bined]] layers.

==== Strand ====
If your input file contains information regarding the strands, you'll be able to choose to load the data from either both or only one strand.

You can also decide to shift the reads from both strands as shown in the figure on the left. To shift the strands just put a value in the "Shift" input box.

The value you entered is going to be added to the position of the data on the 5' strand and subtracted from the ones on the 3' strand.

==== Fragment Length ====
==== Selected Chromosomes ====
By default all the chromosomes of the project are selected. If you want to change this selection, click on the "modify selection" button and uncheck the undesired chromosomes. Working on fewer chromosomes will save memory and loading time.

'''Important Note:''' GenPlay can accelerate the loading if you know that your file is sorted by chromosome. If you press Yes when GenPlay asks you if the file is sorted when your file is actually not sorted, the file may load incompletely, leading to a loss of valuable information. The chromosomes must be ordered the same way it is ordered in the chromosome selection combo-box.

==== Examples of Score Calculations ====
===== For non bined layer =====
====== Example 1 ======
''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

''' Result '''
[[image:loadVWT_ex1.png|center|frame|Loading of an alignment file as a variable window layer]]

----

====== Example 2 ======
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadVWT_ex2.png|center|thumb|400px|Loading of an interval file as a variable window layer]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1010
| 1020
| 100
| 100
| 100
|-
| Chr1
| 1020
| 1120
| (100 + 30) / 2 = 65
| Max(100, 30) = 100
| 100 + 30 = 130
|-
| Chr1
| 1120
| 1300
| (100 + 120) / 2 = 110
| Max(100, 120) = 120
| 100 + 120 = 220
|-
| Chr1
| 1300
| 1350
| 100
| 100
| 100
|}

===== For bined layer =====
====== Example 1 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score is always one)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1135
| 1136
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 1
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex1.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 1
| 1
| 5
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 1
| 1
| 2
|}

----

====== Example 2 ======
Loading of an alignment file as a fixed window layer with a window size of 100:

(each line represents one read position, score varies)

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1125
| 1126
| 1
|-
| Chr1
| 1135
| 1136
| 3
|-
| Chr1
| 1145
| 1146
| 1
|-
| Chr1
| 1149
| 1150
| 1
|-
| Chr1
| 1175
| 1176
| 1
|-
| Chr1
| 1210
| 1211
| 1
|-
| Chr1
| 1230
| 1231
| 1
|-
| Chr1
| 1340
| 1341
| 6
|-
| Chr1
| 1345
| 1346
| 1
|}

[[image:loadFWT_ex2.png|center|frame|Loading of an alignment file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| 7 / 5 = 1.4
| 3
| 7
|-
| Chr1
| 1100
| 1200
| 1
| 1
| 2
|-
| Chr1
| 1200
| 1300
| 7 / 2 = 3.5
| 6
| 7
|}

----

====== Example 3 ======
Loading of an interval file as a fixed window layer with a window size of 100:

''' Input file '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Score
|-
| Chr1
| 1020
| 1120
| 30
|-
| Chr1
| 1120
| 1300
| 120
|-
| Chr1
| 1010
| 1350
| 100
|}

[[image:loadFWT_ex3.png|center|frame|Loading of an interval file as a fixed window layer with a window size of 100]]

''' Result '''
{| cellpadding="4" cellspacing="0" border="1"
! Chr
! Start
! Stop
! Average
! Maximum
! Sum
|-
| Chr1
| 1000
| 1100
| (26.47 + 24) / 2 = 25.23
| Max(26.47, 24) = 26.47
| 26.47 + 24 = 50.47
|-
| Chr1
| 1100
| 1200
| (29.41 + 6 + 60) / 3 = 31.80
| Max(29.41, 6, 60) = 60
| 29.41 + 6 + 60 = 95.41
|-
| Chr1
| 1200
| 1300
| (29.41 + 60) / 2 = 44.70
| Max(29.41 +60) = 60
| 29.41 +60 = 89.41
|-
| Chr1
| 1300
| 1400
| 14.70
| 14.70
| 14.70
|}
 
=== Loading a Gene Annotation Layer ===
[[image:gene_track.png|left|thumb||A Gene Layer]]
[[image:score_color.png|right|thumb|40px|Score Color]]
Select the “Gene Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a gene layer.

Once it's done, just wait until the loading is complete and the gene layer will appear in the track you selected.

Note that the genes on the plus strand are in red and the genes on the minus strand are in blue. If the file contains expression values, the exons are color coded to represent the expression (red = high, blue = low, as shown on the right).
 
=== Loading a Repeat Family Layer ===
Select the "Repeat Layer" option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a repeat layer.

This layer type displays repeats organized by family or class.
 
=== Loading a DNA Sequence Layer ===
Select the “DNA Sequence Layer” option. This opens up a file chooser dialog box that allows you to select the file that you want to load. Please refer to the [[#File formats|File formats]] section if you want to know what kind of file can be loaded as a sequence layer.
[[image:sequence_track.png|center|thumb|300px|A Sequence Layer]]
Sequence layers show DNA sequences from .2bit files.

The hg18, hg19, mm8 and mm9 sequence files can be downloaded from the [http://129.98.70.162/wiki/index.php/Library library] of GenPlay.
 
=== Loading a Mask Layer ===
[[image:Stripes.png|right|thumb|200px|CPG Islands Shown As Stripes On a Refseq Gene Layer]]
Select the "Mask Layer" option. The stripes acting as masks can be useful to show regions of interest such as CpG Islands or repeat regions.

Check the [[#File Formats|File Formats]] section out if you need to know what kind of file can be loaded as a stripes.
 
=== Loading a Variant Layer ===
[[image:Mg_add_layer_variant_selection.png|right|thumb|200px|Add a Variant Layer]]
Select the "Variant Layer" option, this option is only available in multi-genome projects. This will pop up a new dialog to select which sample the user wants to load, and which variation(s).
A variant layer is according to only one sample. It is also possible to change the colors of each variation independently by clicking on the colored square next to the variation checkbox.
 
=== Loading Data From a DAS Server ===
The distributed annotation system (DAS) is a client-server system in which a client can retrieve data from one or multiple servers. GenPlay can connect to any server that follows the DAS/1 protocol as specified by [http://www.biodas.org/wiki/DAS/1 BioDAS]

[[image:DAS_dialog.png|left|thumb||DAS Dialog]]
The “Add Layer from DAS Server” option from the track handler menu will show the DAS Dialog.

Select the server from which you want to retrieve the data in the "Server" box.

Then select the "Data Source". Most of the time, the Data Source corresponds to the reference genome that you want to work on.

Once that's done you need to select the data that you want to retrieve in the "Data Type" box.

GenPlay can either generate a gene layer or a variable window layer from the retrieved data. You can select what type of output layer you want in the "Generate" option.

Finally, you can also choose to download data on only a part of the genome. This can be useful because retrieving data from a DAS server can be time consuming.

'''Note:''' The [[#DAS server|DAS server]] section shows how to add new servers to the list of available servers in the DAS dialog.
 
== Main Menu ==
[[image:main_menu.png|right|thumb|200px|Main Menu]]
On GenPlay’s main screen, click on the top left button (shown by a little hammer and wrench) to pop up the main menu.

=== New Project ===
This will pop up the welcome screen in order to start a new project. All work not saved will be lost.
 
=== Load / Save Project ===
This menu allows you to load or to save a whole GenPlay project in a space efficient binary compressed format. When you load a GenPlay project, all the tracks and layers of your current project will be replaced by the ones from the loaded project and all the information that hasn't been saved will be lost.
'''Important Note:''' The GenPlay project files may be dependent on the version of GenPlay you're using. Be sure to remember with which version of GenPlay you saved a project and use the same version next time you load your project.
 
=== Full Screen ===
Click on this item from the main menu to toggle the full screen mode. When the full screen mode is on, the control panel and the status bar are hidden.

You can also toggle the full screen mode by pressing the F11 key.
 
=== Warnings report ===
This option will pop up the Warnings report dialog in order to consult previous and current alerts.
 
=== Option ===
The option menu item allows you to modify the configuration of GenPlay. Please refer to the section [[#Changing the Configuration of GenPlay|Changing the configuration of GenPlay]] for further information.
 
=== RNA To DNA Reference ===
This option allows you to transformed the coordinate system of the result of a RNA-Seq experiment based on alignment to a transcriptome (for instance all refseq genes), to a genomic coordinate system.

You need two files in order to use this functionality.
# The result of the RNA-Seq experiment, called "Coverage File" in GenPlay. This file must be in bedGraph file format.
# An annotation file in bed format.

Two output files can be generated:
# A bedGraph file with the position based on a reference genome
# A annotation GdpGene file

Here is an example:
Coverage File:
NM_000016 0 413 0
NM_000016 413 456 1
NM_000016 456 471 2
NM_000016 471 488 3
NM_000016 488 494 2
NM_000016 494 504 3

Annotation File:
chr1 76190042 76229353 NM_000016 0 + 76190472 76228448 0 12 460,88,98,70,101,81,131,109,141,96,249,977, 0,4043,8286,8495,9170,10433,15622,21448,25061,26093,36764,38334,

The result as a bedGraph file is:
chr1 76190455 76190498 43.0
chr1 76190498 76190502 8.0
chr1 76194085 76194096 22.0
chr1 76194096 76194113 51.0
chr1 76194113 76194119 12.0
chr1 76194119 76194129 30.0

And the result as a GdpGene file is:
NM_000016 chr1 + 76190042 76229353 76190042,76194085,76198328,76198537,76199212,76200475,76205664,76211490,76215103,76216135,76226806,76228376 76190502,76194173,76198426,76198607,76199313,76200556,76205795,76211599,76215244,76216231,76227055,76229353 667888.95,1506024.1,0,0,0,0,0,0,0,0,0,0
 
=== Help and About GenPlay ===
The help and the about GenPlay options open a browser showing respectively the documentation and about pages of GenPlay website.
 
=== Exit ===
This option closes the application after asking for confirmation.
 
== Changing the Configuration of GenPlay ==
[[image:main_menu.png|right|thumb|250px|Option Menu]]
Click on the option item of the [[#Main Menu|main menu]] to open the configuration screen.
 
=== General Options ===
The following screen lets you set the general options.

The Default Directory lets the user choose which folder to open by default for any of the file chooser within GenPlay.

From this screen, you can also modify the appearance of the software by changing the look & feel.
 
=== Track Option ===
The Number of Tracks text box defines the maximum number of tracks that can be loaded on GenPlay.

The Default Track Height text box defines the height of each of the tracks.

The Undo Count text box defines the number of operations that can be undone. Note that the higher the number of undos selected, the more memory will be required.

The reset option allows the user to easily reset a layer in order to come back as if it has been freshly loaded.

The legend showing layers name on the upper right of a track can also be enabled or disabled.
 
=== DAS Server ===
The DAS server option shows the list of existing DAS servers along with the URL where these servers are located. It also provides the options to add new servers and remove existing servers.

GenPlay can communicate and retrieve data from the servers implementing the [http://www.biodas.org/wiki/DAS/1 DAS/1 protocol]
 
=== Restore Default ===
The Restore Default configuration restores everything back to the factory settings.
<gallery widths=220px heights=100px perrow=3>
image:Options_general.png|General Options
image:options_track.png|Track Option
image:das_server_option.png|DAS Server
</gallery>
 
== File Formats ==
The different file formats used in GenPlay are described on this [[GenPlay File Formats|page]].
 
== Using Tracks ==
[[image:add_layer.png|right|thumb|150px|Track Menu]]
=== Handling Tracks ===
==== Moving a Track ====
To move a track up or down in the track list, just click on the track handler (the left part of the track with the track number) and drag the track to the desired position.
 
 
 
==== Inserting a Track ====
To insert a track, right click on the track handler of the track right under where you want to insert and choose the "Insert" option.
 
 
 
==== Deleting a Track ====
To delete, select a track and click on the delete option of the contextual menu or press Delete on the keyboard.
 
 
 
==== Copying, Cutting and Pasting a Layer ====
[[image:paste_layer.png|right|thumb|200px|Track Menu]]
To copy layers, select the desired track where the layers are and click on the copy option in the contextual menu or press CTRL+C.

To cut layers, select the desired track where the layers are and click on the cut option in the contextual menu or press CTRL+X.

To paste a track, select the track where you want to paste and click on the paste option in the contextual menu or press CTRL+P. 
A new window will appear showing all layers recently copied/cut that can be pasted on the track. The user has to select all layers he wants to paste and then click "Ok".
 
 
 
==== Taking a Screenshot of the Track ====
To take a screenshot, select a track and choose the "Save as Image" option in the contextual menu.
 
 
 
==== Using the Undo / Redo / Reset Options ====
The undo, redo and reset options are only available for the Variable and Fixed Window layers. They are accessible from the contextual menu when you right click on the track handler.

The number of undo and redo operations available can be specified as described in the [[#Track Option|Track Option]] section. Note that this operations are memory consuming and reducing the number of undo / redo available can save memory.

The reset operation restore the track to the way it was right after being loaded. A reset operation can also be undone.
 
=== Track/Layer Settings ===
==== General ====
[[image:track_settings_track.png|right|thumb|300px|Track Settings - General]]
===== Basic Options =====
*Name: The name of the track.
*Height: The height of the track.
===== Axis Options =====
*Show horizontal lines: Split the track horizontally.
*Horizontal line count: Number of horizontal lines, equally separated.
*Show vertical lines: Split the track vertically.
*Vertical line count: Number of vertical lines, equally separated.
===== Score Options =====
*Minimum Score: The minimum score to show.
*Maximum Score: The maximum score to show.
*Auto-rescaled: Enable the automatic score rescaling.
*Score Position: Choose where the score is shown (top/bottom).
*Score Color: Set the font color of the score.
==== Layers ====
[[image:track_settings_layer.png|right|thumb|300px|Track Settings - Layers]]
*Name: Click on the name to edit it.
*Type: The type of layer.
*Color: Click to edit the color of the layer.
*Graph Type: Click to change the graph type:
**Curve
**Points
**Bar
**Dense
*Visible: Show/hide the layer.
*Active: Set the layer as "active". The active layer as direct interaction with the mouse pointer and clicks.
*Set For Deletion: If set, the layer(s) will be deleted when clicking "Ok".
 
== Operations ==
Once a layer is loaded, a right click on the location of the track handler opens a popup menu as shown in the figure below.
[[image:operation_menu.png|center|thumb|600px|Operation Menu]]
The Operation sub-menu of the popup menu contains all the actions that you can use on the selected layer.
=== Sequencing/Microarray Layer Operations ===
Bin-ed and non bin-ed layers do not have all the same operations. They share most of them but some are specific.
<gallery widths=250px heights=400px perrow=2>
image:micro_seq_operations.png|Non bin-ed Microarray/Sequencing Layer Operations
image:micro_seq_bin_operations.png|Bin-ed Microarray/Sequencing Layer Operations
</gallery>
==== Common operations ====
===== Show History =====
Show the history of the layer, every changes that have been made since loaded.
===== Constant Operation =====
[[image:Micro_seq_constant_operation.png|right|thumb|250px|Operation With Constant]]
Thes operations use one constant in the following ways:
* Addition: adds the constant to each window (F(x) = x + constant).
* Subtraction: substracts the constant to each window (F(x) = x - constant).
* Multiplication: multiplies the score by the constant(F(x) = x * constant).
* Division: divides the score by the constant (F(x) = x / constant).
* Inversion: inverts the score of each windows (F(x) = constant / x).
* Unique Score: sets all windows to an unique score (F(x) = constant).
The function can also be applied to null windows by checking the box.
===== Two Layers Operation =====
This allows operations between two Sequencing/Microarray layers, bin-ed and non bin-ed. 
In order to set the operations, few windows appear in the following order:
# A first window appears in order to select the second layer.
# The second window asks in which track the resulting layer will be put.
# The third and last window offers the algorithms to complete the operation (x1: score first layer; x2: score second layer):
* Addition: add scores (x = x1 + x2).
* Subtraction: substract scores (x = x1 - x2).
* Multiplication: multiply scores (x = x1 * x2).
* Division: divide scores (x = x1 / x2).
* Average: average score (x = (x1 + x2) / 2).
* Maximum: keeps the highest score.
* Minimum: keeps the lowest score.

'''Note:''' The only way the resulting layer would be a bin-ed layer is to make an operation between two bin-ed layer having the same bin size. Any other case will result in a non bin-ed layer.
===== Index =====
Indexation can be useful to compare multiple layers at the same scale. It "re-scales" existing scores to a new range defined by the user. 
If scores go from 10 to 600 but for some reason would need to be observed between 0 and 100, this operation will do the work. 
It will first ask for the new minimum and the new maximum. The next dialog asks to perfom the re-scaling by chromosome independently or genome wide. 
Using the previous example, for a new scale of [0; 100] if the first chromosome as a maximum score of 600 and the second one has a maximum score of 800; 800 will become the reference value of 100 for both chromosomes if the operation is processed genome wide. If the operation is processed by chromosome independently, 600 will become the reference value of 100 for the first chromosome, and 800 for the second chromosome.

Since this operation uses the minimum and maximum scores, it is very important to note that indexing does not work well in the presence of outliers. Indexing works best if outliers are eliminated or removed first using a filter (see below).
===== Log =====
[[image:operation_log.png|right|thumb|100px|Logarithm Bases]]
For each window, the log operation applies the function f(x) = log(x), where x is the window score. The base of the logarithm function can be selected between either 2 (binary log), e (natural log) or 10 (common log).
===== Normalize =====
[[image:operation_Normalize.png|left|thumb|100px|Normalization Coefficient]]
After a normalize operation the score of each window is divided by the result of the Score Count operation and multiplied by a specified fixed value. By default, after normalization the scores are expressed per 10 millions reads.
===== Standard Score =====
Calculates the standard score for the selected layer i.e. (x - avg) / stdev; where x is the score, avg is the average score of the layer and stdev is the standard deviation of the scores of the layer.
===== Filter =====
GenPlay provides four different filters:
====== Percentage Filter ======
[[image:operation_pfilter.png|right|thumb|130px|Percentage Filter]]
This option filters the X% lowest values and the Y% greatest values where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Threshold Filter ======
[[image:operation_tfilter.png|right|thumb|130px|Threshold Filter]]
This option removes the values that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
====== Band-Stop Filter ======
[[image:operation_bfilter.png|right|thumb|130px|Band-Stop Filter]]
This option removes values between two specified threshold.
====== Count Filter ======
[[image:operation_cfilter.png|right|thumb|130px|Count Filter]]
This option filters the X lowest values and the Y greatest values, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Transfrag =====
This operation aggregates the windows of the selected layer that are separated by a gap smaller than a specified size (in bp).

The score of the new window can be the sum, the average or the maximum of the scores of the aggregated windows.
===== Score Distribution Histogram =====
The show repartition operation generates a graph showing the distribution of the scores of the selected layers. The options for the type of plot are score v/s window count and score v/s base pair count.

The user needs to choose a size for the bins of scores. The graphics will show, depending on the selection, how many windows or how many base pair there is for each bin of scores.
===== Convert Layer =====
This operation converts the current layer into another layer among the following:
*Gene Annotation Layer
*Microarray/Sequencing Layer bin/non-bin
*Mask Layer
==== Non Bin-ed Layers Only ====
===== CG Methylation Profile =====
This operation computes the methylation values on CG sequences by combining the value on the C position and the value on the G position. 
This is based on data fron a sequence layer in order to find CG sequences.

==== Bin-ed Layers Only ====
===== Smooth =====
The smooth operation can be processed according to the 3 following algorithms:
====== Gauss Smoothing ======
[[image:operation_fwt_gaussian.png|right|thumb|150px|Sigma Value]]
This operation applies a [http://en.wikipedia.org/wiki/Gaussian_filter Gaussian filter] to the layer, depending on the sigma value provided by the user.

G(x) = (1 / v (2?) s) * e-x2 / 2 s2

Where, x is the score and s is the standard deviation of the layer.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Loess Smoothing ======
This operation computes the Loess regression of degree 1 on the selected layer.

For each x value where a y value is to be calculated, the Loess technique performs a regression on points in a moving range around the x value, where the values in the moving range are weighted according to their distance from this X value.

The Loess regression is a smoothing function. You will need to precise the half size of the moving window on which the regression will be computed.

The weight function of the Loess regression is computed as follow: W(i) = (1 - X(i)^3)^3, where X(i) is the normalized distance: current distance / maximum distance among points in the moving regression.

You can choose the extrapolate option to "fill" the windows with a score of zero.
====== Moving Average Smoothing ======
For each window of the layer, compute the average on a region of a specified size center on the window and score the window with the result of this average. The half-size of the region is prompted prior to the calculation.

You can choose the extrapolate option to "fill" the windows with a score of zero.
===== Find Peaks =====
The find peak operation offers three different algorithms that can be used to find the peaks:
====== Standard Deviation Peak Finder ======
[[image:operation_fwt_sfinder.png|left|thumb|150px|Standard Deviation Peak Finder]]
The standard deviation peak finder prompts the user to enter two parameters.

The parameter ‘S’ specifies the number of windows to be considered for each window on either side in order to calculate the standard deviation.

For example, if S = 10, it means that for each window we consider 10 windows to the left and 10 windows to the right to calculate the standard deviation.

For a window to be accepted, its standard deviation needs to be at least ‘T’ times greater than the value of the standard deviation of the chromosome.
====== Density Peak Finder ======
[[image:operation_fwt_dfinder.png|right|thumb|150px|Density Peak Finder]]
The Density Finder works as follows:

The parameter ‘S’ specifies the number of windows to be considered for each window on either side of the window under consideration.

For the window under consideration to be accepted, at least ‘P’ percentage of values must be above the high threshold ‘H’ or at least ‘P’ percentage of values must be below the low threshold ‘L’.
====== Island Finder ======
[[image:operation_fwt_ifinder.png|left|thumb|150px|Island Finder]]
The Island Finder is based on the algorithm described in the paper
[http://bioinformatics.oxfordjournals.org/content/25/15/1952.full.pdf Zang, C., Schones, D. E., Zeng, C., Cui, K., Zhao, K., and Peng, W. (2009). A clustering approach for identification of enriched domains from histone modification chip-seq data. Bioinformatics (Oxford, England), 25(15):1952-1958.]

The parameters window value and gap of the island finder are the parameters ‘l0’ and ‘g’ respectively. The island score allows the user to select the scores greater than or equal to a particular value. The island length parameter allows the user to select islands encompassing at least specified number of windows.
There are two result types:
* Start values: Depicts only those islands that are selected and removes the ones that are rejected.
* Island score: Depicts the islands by considering the score.
* Island Summit: Depicts the island with the summit of the input island as a score.
===== Correlation =====
[[image:operation_fwt_correlation.png|right|thumb|100px|Correlation Report]]
The correlation operation computes the Pearson’s correlation between the score values of two layers. The two layers need to have the same bin size. The following formula is used to calculate the correlation:

? = ( ? xi yi – n x’ y’) / ((n - 1) sx sy)

Where:
* ? is the Pearson’s correlation
* xi and yi are the scores of the layers
* n is the number of values
* x’ and y’ are the means of the scores of the layers
* sx and sy are the standard deviations of the scores of the layers

The figure on the right shows a correlation report.

'''Note:''' The correlation is computed only on the windows that are different from zero on both layer. If one of the layer has a zero value window, the window of the other layer with the same coordinate will be skipped as well.
===== Density =====
This operation generates a new fixed window layer where the score of the windows represent the density of non null windows in the neighborhood of the windows.
You first need to enter the size S of the neighborhood.
For each window W, the algorithm count how many of the S windows before W and the S windows after W have a score different from zero. This value is then divided by 2 * S + 1 and the result is the score of W.
===== Intervals Scoring =====
This operation needs two layers:
* The selected layer that defines the scores
* A second layer that defines the intervals
This operation generates a new layer containing the intervals of the "interval track". For each interval the algorithm then looks at the corresponding scores in the score layer, and compute either the maximum, the average or the sum of all the scores that fall in the interval. This value is the new score value in the result layer.

You can also choose to use only a certain percentage of the greatest scores that falls in the interval.
===== Concatenate =====
[[image:operation_select_tracks.png|right|thumb|150px|Select Layers to Concatenate]]
The concatenate operations allows you to generate a file containing the scores of multiple fixed window layers that have the same bin size.
The output file contains the following fields:
# chromosome
# start position
# stop position
# score layer 1
# score layer 2
# score layer 3
# ...
 
=== Gene Layer Operations ===
Directly on a gene layer, you can:
# Double click on a gene to open a web page describing the gene. Make sure that your input file contains a geneDBURL line as described in the [[#File Formats|File Formats]] section in order to enable this option.
# Put the mouse over a gene to have some information about the name and the score of the gene. If the exons of the gene have different scores you can put your mouse over an exon to have the exon score.
==== Score Count ====
This operation count the sum of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Average ====
This operation computes the average of all scores. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes ====
This operation count the total number of genes. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Genes with Non-Null Score ====
This operation count the total number of genes excluding the ones with a score of 0. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Count Exons ====
This operation count the total number of exons. 
A window asks first to select chromosomes to include in the calculation (all by default).
==== Search Gene ====
[[image:gene_search_gene.png|left|thumb|100px|Search Gene]]
Use this option to search a gene on the selected layer by typing the name of the gene.

Check the Match Case option if you want the search to be case sensitive.
Check the whole word option if you want to search genes where the input match the whole name of the gene.
Press next or previous to find respectively the next or previous gene found.
You can also open the Find Gene dialog by pressing CTRL+F after selecting a gene layer.
==== Extract Intervals ====
[[image:gene_extract_intervals.png|right|thumb|200px|Extract Intervals]]
This option allows you to extract intervals defined relatively to the beginning, the end or the middle of a gene and to generate a new gene layer showing these intervals.

You can, for example, defined promoters as regions that starts 100bp before the beginning of genes and that ends 150bp after the beginning of genes. This option would allow you to generate a new layer from this parameters.
==== Extract Exons ====
[[image:gene_extract_exons.png|right|thumb|150px|Extract Exons]]
This option generate a new gene layer showing only the exons of the genes of the selected layer.

You can choose between the three following options:
# Extract the first exon of the genes
# Extract the last exon
# Extract all the exons
==== Unique Score ====
[[image:gene_unique_score.png|right|thumb|150px|Unique Score]]
This operation sets the same score for all exons.
==== Score Exons ====
[[image:gene_score_exons.png|right|thumb|200px|Score Exons]]
To execute this operation you need to have at least one microarray/sequencing layer loaded. For each exon of each gene of the selected gene layer, this operation computes a new score based on the window score from the selected layer that falls into the exon. There are 3 different ways to compute the new score:
*Base Coverage Sum
*Maximum coverage
*RPKM
==== Filter ====
This option provides four different filters for gene layers:
===== Percentage Filter =====
[[image:gene_filter_percentage.png|right|thumb|130px|Percentage Filter]]
This option filters the genes with the X% lowest overall score and the Y% greatest overall scores where X and Y are two decimals and where X + Y <= 100.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Threshold Filter =====
[[image:gene_filter_threshold.png|right|thumb|130px|Threshold Filter]]
This option filters the genes with an overall score that are lower than X OR greater than Y, where X and Y are two specified threshold values.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
===== Band-Stop Filter =====
[[image:gene_filter_band-stop.png|right|thumb|130px|Band-Stop Filter]]
This option removes the genes with an overall score between two specified threshold.
===== Count Filter =====
[[image:gene_filter_count.png|right|thumb|130px|Count Filter]]
This option filters the X lowest scored genes and the Y greatest scored genes, where X and Y are two specified integers.
You can choose between removing the filtered values (remove) or setting the filtered values to the boundary values (saturate).
==== Filter Strand ====
You need to select a strand when prompted. At the end of the operation the layer will contain only the genes on the selected strand. All the other genes will have been removed.
==== Rename Genes ====
This operation allows you to change the name of the genes. You need to provide a text file where each line contains the current gene name and the new gene name separated by a tabulation. Every time a gene with a name from the first column is found this name will be replace by the new gene name from the second column.
==== Distance Calculation ====
Development in progress, coming soon.
==== Score Repartition Around Start ====
You first need to select a Fixed window layer containing the scores. After that, you need to select the chromosomes on which you want to execute the operation. You also need to specify a bin size S, a bin count C and a method for the calculation of the scores.

The operation will create C bins on each side of the start position of each gene. The size S of each bin is in base-pair. Depending of the method of calculation chosen the operation is going to compute the sum, the maximum or the average of the scores for each corresponding bin from each gene and display a bar graph of the result. The data can be exported by right-clicking on the graph and using the "save as" function.

Multi-curve graph can be generated using the following procedure: 
To generate a comparison between 2 fixed-window layers: 1) Perform an analysis for the first layer as described above. 2) Save it to your hard drive. 3) Close the graph window. 4) Perform the same analysis on the second layer. 4) Right click on the second graph and choose the load data option. 5) Load the first analysis. Colors of the curves, type of graphs (bar, points, curve) and scale can be adjusted by right-clicking on the graph. Procedure can be used to load more than two graphs. To produce more complex graphs we recommend loading the saved data on your favorites spreadsheet software.
Score Repartition Around Start
 
=== Repeat Layer Operations ===
There is currently no operation available for the repeat layer.
 
=== DNA Sequence Layer Operations ===
There is currently no operation available for the sequence layers.
 
=== Mask Layer Operations ===
Documentation in writing...
 
=== Variant Layer Operations ===
Documentation in writing...

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