The results of differential gene expression analyses (GSA or ANOVA) in Partek Flow can be filtered to extract meaningful results, e.g. statistically significant genes. User-defined filters can be saved and applied to other projects.
On a Feature list task report, specify the criteria to filter the table of statistical results using the Filter dialog (Figure 1) and click the Save filter button.
Name the filter by typing into the Filter name box in the Save filter dialog and click the Save button (Figure 2).
Saved filters will appear in the Saved filters section in the Filter dialog (Figure 3). Saved filters are only available to the Partek Flow user that created them. They will not be available to other administrative or non-administrative users on the same Partek Flow instance.
To apply a saved filter to a table of statistical results, click the filter name in the saved filters section in the Filter dialog. If a filter is applied to a table, the filter name will appear at the top of the Filter dialog.
Hovering over the Preview filter () icon in the Filter dialog will show a full description of the filter in a pop-out balloon. Applied filters are displayed at the top of the Filter dialog
Filter Management Page
To access the Filter management page, click on the gear () icon in the Filter dialog on a Feature list task report (Figure 1, above). Alternatively, click the avatar in the top right corner, choose Settings and click Filters on the left (Figure 4). This page can be accessed by administrative and non-administrative Partek Flow users. The table on the Filters page displays all saved filters created by the Partek Flow user that is currently logged in (Figure 4). Filters created by other Partek Flow users will not be displayed in the table.
Click the icon in the Actions column to show actions available for the filter (Figure 5).
From the action list, select View filter details to show the filter details (Figure 6). To delete a filter, click the Delete filter button (Figure 5)
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Selecting an Assembly
Choosing an Assembly from the drop-down list at the top will display the associated library files in the panel below. Assemblies are named by the species and the build version (e.g. Homo sapiens (human) - hg19). Different build versions for the same species are regarded as separate assemblies (Figure 1). Administrative users can delete a selected assembly by clicking the bin () icon.
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Figure 1. Filter dialog showing filtering criteria (false discovery rate (FDR) step up ≤ 0.05 & Fold change <-2 or >2) and indicating no filters have been saved yet. The Save filter button is at the bottom of the dialog. The gear icon opens the Filter management page
Figure 2. Give the filter a name and save
Figure 3. Saved filters are available at the bottom of the Filter dialog
Figure 4. The Filter management page
Figure 5. Actions that can be performed on a filter
Figure 1. Human assembly versions hg18, hg19 and hg38 will appear as separate assemblies in the drop-down list.
Adding a Cytoband
Click the + Add reference files button under the Reference Files section header and choose Cytoband from the Library type drop-down list in the Add reference sequence dialog (Figure 1). If a reference sequence is already associated with an assembly, Cytoband will be the only option available in the Add reference sequence dialog and will not appear in a drop-down list (Figure 1).
Figure 1. Add reference sequence dialog. If the reference sequence and cytoband files have not been added yet, both options will appear in the Library type drop-down list (top). If one is missing, it will appear as the only option (bottom). For many model organisms, automatic downloads are available from the Partek repository
If you are using an assembly supported by Partek (e.g. human), select Download cytoband and click Create to get the cytoband file from the Partek repository. Alternatively, select Build cytoband and select .fasta files or .2bit files to to build the cytoband file. If the reference sequence is missing, it will either be downloaded automatically or you will be asked to import it from another source (see Adding a reference sequence).
If you are using a custom assembly (e.g. for a non-model organism), the Download cytoband option will not be available (Figure 2).
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Creating an Assembly on the Library File Management Page
On the library file management page, an assembly can be added by choosing Add assembly… from the Assembly drop-down list (Figure 1). If the list is long, you may need to scroll to the bottom.
Figure 1. Adding an assembly on the library file management page
In the Add assembly dialog, choose the Species from the drop-down list (Figure 2), followed by the Assembly version (Figure 3) and click Add. The dialog will automatically load commonly used assembly versions for the selected species (Figure 3). If the assembly version you want does not appear in the list, choose Other and type the custom assembly version name (Figure 4). Characters such as $ * | \ : " < > ? / % cannot be used in custom names. Note that If an assembly version for a given species already exists on your system, it will not appear in the drop down list.
Figure 2. Add assembly dialog. The species can be chosen from the drop-down list
Figure 3. For the chosen species, commonly used assembly versions can be chosen from the drop-down list
Figure 4. A new assembly version can be added by choosing Other and typing the custom assembly version name manually
If the species you want to add does not appear in the list, scroll to the bottom of the species list, choose Other and manually type the species name and assembly version (Figure 5).
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Update Library Index
The library index is a file that contains download sources for all Partek distributed library files. An automatic update of this file will occur every 24 hours (Figure 1), so this normally doesn't require any attention. If you do wish to manually update the library index, click Update Library Index at the bottom of the Library file management page.
Figure 1. The date and time the library index was last updated is displayed at the bottom of the library file management page
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Adding a Reference Sequence
Click the + Add reference files button under the and choose Reference sequence from the Library type drop-down list in the Add reference sequence dialog (Figure 1). If a cytoband file is already associated with an assembly, Reference sequence will be the only option available in the Add reference sequence dialog and will not appear in a drop-down list (Figure 1).
If you are using an assembly supported by Partek (e.g. human), select Download reference sequence and click Create to get the reference sequence from the Partek repository. Alternatively, select Import reference sequence and click Create to add the reference sequence from another source.
If you are using a custom assembly (e.g. for a non-model organism), the Download reference sequence
Library File Management Settings
To review and edit library file management settings, you must be logged into Partek Flow as a user with administrator privileges. Click on the avatar in the top right corner and choose Settings. Then click System preferences on the left.
You can review the current library file directory location (Figure 1).
To change where the library files are stored, click Edit filesystem and storage and click Browse () icon to point to another directory (Figure 2).
Library File Management Page
To access the library file management page click the avatar in the top right corner and choose Settings. Then under Components on the left, click Library files.
The library files page has three tabs - Genomic library files, Prep kits files, and Other library files. This section of the user guide will focus on the Genomic library files tab, which is relevant for next-generation sequencing analysis (Figure 1).
The arrows ( v / ) expand/collapse each section. Associated library files are shown in a table in each section. In the Actions column of each table, the View library file details button displays additional library file details, the bin () icon dissociates a library file. The hourglass () icon indicates a library file is being created.
Microarray Library Files
Microarray library files can be managed under Microarray library files section on Other library files tab on the Library file management page (Figure 1). The chip name and download source of stored Microarray library files will be shown in the table in this section. For more information, refer to the section.
Microarray probe tab files are used for processing microarray data in Partek Flow. When microarray intensity data files (e.g. Affymetrix .CEL files) are imported into a project, the chip type is automatically detected and the appropriate probe tab annotation file is downloaded. Thus, you would normally not need to manually add any probe tab annotation files.
To add a (e.g. for a custom chip), click the + Add probe sequence button at the Microarray library files section (Figure 1). Scroll to the bottom of the Chip name drop-down list and choose Other / Custom. Name the chip by typing into the Custom name box and click the Create button (Figure 2). Characters such as $ * | \ : " < > ? / % cannot be used in custom names.
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Figure 1. System preferences page showing library file directory location and privilege settings. Settings can be changed by clicking on Edit button at each section
Figure 2. Changing where the library files are stored
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Figure 5. A new species can be added by typing the name and assembly name manually
option will not be available (Figure 2).
Figure 2. For custom assemblies, automatic downloads are not available so the reference sequence has to be imported from another source
The Import reference sequence option allows you to add a reference sequence from the Partek Flow Server or a URL download link (Figure 3).
Figure 3. A reference sequence can be added from multiple sources
To add a reference sequence from the Partek Flow server, select Partek Flow Server, click Browse icon and navigate to the directory storing the reference sequence file(s) (Figure 4). Only files with the correct file extensions (.fa, .2bit or any compressed format) will be displayed in the file browser. Select the checkbox next to the file name(s) you wish to import and click Continue (Figure 4). Click Finish to add the selected reference sequence file(s).
Figure 4. Add a reference sequence file(s) from the Partek Flow Server
To upload a reference sequence from your local computer, click Transfer files to the server, click Transfer files, drop files to the Transfer files dialog or click My Device and navigate to the location of the file(s) on your local machine (Figure 5). Click Upload to complete the file upload. Do not exit the browser tab or let the computer go to sleep or shut down until the transfer has completed.
Figure 5. Upload a reference sequence file from your local computer through the browser
To download a reference sequence (e.g. from a public repository such as ENSEMBL), select URL and input the URL address (Figure 6). Make sure to include the full URL address, starting with the transfer protocol (e.g. http:// or ftp://) and ending with the file name. It is advisable to find the download link address, copy it to the clipboard and paste it into the Input URL box. Click Finish to add the selected reference sequence file(s) (Figure 6).
Figure 6. Download a reference sequence from the internet
Note that you can specify multiple fasta files (e.g. one file per chromosome) and Partek Flow will concatenate them. If a fasta file is imported, a 2bit version of the file will automatically be created and vice-versa. Partek Flow will also accept a range of compressed file formats (gzip, bzip2, zip, tar). If a compressed file is imported, it will automatically be uncompressed and processed.
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Figure 1. Add reference sequence dialog. If the reference sequence and cytoband files have not been added yet, both options will appear in the Library type drop-down list (top). If one is missing, it will appear as the only option (bottom). For many model organisms, automatic downloads are available from the Partek repository.
Figure 2. Adding a custom probe tab file e.g. for a custom microarray chip
A custom probe tab file can be added from the Partek Flow Server or a URL download link. The behavior of each option is similar to when importing a reference sequence (see Adding a Reference Sequence). When browsing for files on the Partek Flow server, only the files with relevant file extensions will be visible (.probe_tab and various compressed formats). Please see the Importing Custom CEL files user guide for more information.
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Missing library files can be added when setting up tasks within a project, without having to navigate to the library file management page. The user interface will vary depending on the task and which library files already exist on your system. Below are two examples scenarios.
Adding a missing whole genome aligner index (Bowtie 2 in this example)
Under the Analyses tab of a project, select an Unaligned reads data node
From the context sensitive menu on the right, choose Aligners followed by Bowtie 2. On the alignment task setup page, Partek Flow will display all assemblies that have a Bowtie 2 index (whole genome and transcriptome) in the Assembly drop-down list. If the assembly you want is missing, choose New assembly… from the drop-down list (Figure 1)
Choose the species and assembly in the Add Bowtie 2 index dialog. If the species and assembly you want do not appear in the drop-down lists, choose New species... and manually type the names (Figure 2)
Choose Whole genome from the Index drop-down list (Figure 2)
Select the Build index radio button (Figure 2)
Once the new Bowtie 2 index has been specified, you are able to queue the alignment task and it will execute once the Bowtie 2 index has been built.
Add a missing gene set file for enrichment analysis
Under the Analyses tab of a project, select a Feature list data node
Choose Biological interpretation from the menu on the right, followed by Gene set enrichment
Select Gene set database radio button for Database. If the species and assembly you want do not appear in the Assembly drop-down list, select New assembly... (Figure 3)
If you are working with an assembly/species supported by Partek (e.g. human), choose a gene set from the Gene set database drop-down list (Figure 4), select the Download gene set database radio button and select Create. Alternatively, choose Add gene set database from the Gene set database drop-down list, manually type the custom gene set name and click Create to import your own gene set from the Partek Flow server or URL (Figure 5). Characters such as $ * | \ : " < > ? / % cannot be used in custom names. If you are working with a custom species/assembly (e.g. for a non-model organism), only the Add gene ontology source option is available.
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Library File Management
The library file management tool in Partek Flow provides an easy way to create, process and manage reference sequences, cytoband files, annotation models, aligner indexes, gene sets, variant databases and microarray probe sequence files.
For a quick start, please watch the short videos below:
Quickly setup library files for the hg19 human reference genome assembly.
The process can be adapted to the different kinds of file types.
Adding library files for a non-model organism
You can watch the .
This user guide will cover the following topics:
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Adding a Gene Set
Click the + Add gene set databases button under the Gene set databases section header on the library file management page.
If you are using an assembly supported by Partek (e.g. human), a gene set from geneontology.org will appear in the Gene set database drop-down list (Figure 1). Select the Download gene setdatabase radio button and click Create.
Figure 1. For many model organisms, automatic downloads of gene sets are available from the Partek repository (source: http://geneontology.org/)
If you prefer to add a custom gene set, or if you are working with a custom assembly, choose Add gene set database from the Gene set database drop-down list (Figure 2). Name the gene set by typing into the Custom Name box and click Create. Characters such as $ * | \ : " < > ? / % cannot be used in custom names. A gene set file can be added from the Partek Flow Server or a URL download link. The behavior of each option is similar to when importing a reference sequence (see Adding a Reference Sequence). When browsing for files on the Partek Flow server, only the files with relevant file extensions will be visible (.gmt and various compressed formats).
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Adding an Annotation Model
On the library file management page, click the +Add annotation models button in the Annotation models section, choose Gene/feature annotation from the Library type drop-down list in the dialog (Figure 1).
If you are using an assembly supported by Partek (e.g. human), annotation models from a variety of commonly used sources (e.g. ENSEMBL, GENCODE) will appear in the Annotation model drop-down list in the dialog. Choose an annotation model, select the Download annotation file radio button and click Create (Figure 1).
Figure 1. For many model organisms, automatic downloads of gene/feature annotation models from various sources are available from the Partek repository.
If you prefer to add a custom annotation file or if you are working with a custom assembly, choose Add annotation model from the Annotation model drop-down list (Figure 2). You may need to scroll to the bottom of the drop-down list to see this option. Name the annotation model by typing into the Custom name box and click Create. Characters such as $ * | \ : " < > ? / % cannot be used in custom names.
A custom annotation model can be added from the Partek Flow Server or a URL download link. The behavior of each option is similar to when importing a reference sequence (see ). When browsing for files on the Partek Flow server, only the files with relevant file extensions will be visible (.gtf, .gff, .gff3, .bed, .pannot, .txt and various compressed formats).
Select the source of annotation file and click Next. If the annotation file format is gtf, gff, gff3 or bed, a preview of the first 10 rows of the file will be shown on the screen (Figure 3). You must then specify the type of annotation file by choosing an option from the Annotation type drop-down list (Figure 3). Choose mRNA for whole transcriptome annotations, where both gene and transcript level information are present. Choose microRNA for precursor or mature microRNA transcript annotations. Choose Amplicon for targeted amplicon sequencing annotations. Choose Other for annotation files that do not fall into the other four categories (e.g. lncRNA).
For text annotations with both gene- and transcript-level information select mRNA from the drop-down menu. You can manually specify which column corresponds to the Transcript ID or Gene ID by selecting the corresponding radio button next to the column with that information (Figure 4).
Genome coordinates for annotation models stored in Partek Flow are 1-based, start-inclusive, and stop-exclusive. This means that the first base position starts from one, the start coordinate for a feature is included in the feature and the stop/end coordinate is not included in the feature. These are the genome coordinates that are printed in various task reports and output files when an annotation model is involved in the task. When custom annotation files are added to Partek Flow, the genome coordinates are converted into this format. The coordinates are converted back if necessary for a specific task. Figure 5 shows how the genome coordinates vary between different annotation formats.
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Adding a Variant Effect Predictor (VEP) Database
Click the + Add VEP database button under the VEP database section on the .
If you are using an species supported by Partek (i.e., human, mouse, rat), a VEP database from will be available for automatic download. Select the Download database radio button and click Create (Figure 1).
If you are working with a custom assembly, choose the Import database radio button and click Create (Figure 2). A VEP database can be added from the Partek Flow Server or a URL download link. The behavior of each option is similar to when importing a reference sequence (see ). ENSEMBL maintains an extensive list of VEP databases for various species. We recommend going to and finding the VEP database download link for your species of interest.
Adding a SnpEff Variant Database
Click the + Add SnpEFF variant databases button under the SnpEff variant databases section header on the .
If you are using human (hg19 and hg38), mouse (mm10) or rat (rn5 and rn6) assemblies, various versions of SnpEff variant databases are available for automatic download (Figure 1).
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Pipeline managment
The Pipelines management page allows users to view and manage the pipelines within a Partek Flow instance. To access the page, click the avatar on the top right corner, choose Settings, then select Pipelines on the left. This page can be accessed by administrative and non-administrative users. The table (Figure 1) shows a list of pipelines with their description, creation date, and the display name of the owner (the user who created or imported the pipeline). Non-administrative users can see pipelines created by other Partek Flow users in the table if the pipeline has been shared by the user. Administrative users can see all details.
Hiding Pipelines
Adding Aligner Indexes Based on an Annotation Model
Note that this task is for adding indexes for alignment to a subset of the genome (e.g. the transcriptome). If you want to align to the whole genome, see .
On the , click the + Add annotation models button at the Annotation models section and choose Aligner index from the Library type drop-down list in the dialog (Figure 1).
Choose the aligner you wish to use from the Aligner drop-down list (Figure 1). All aligners are available for indexing to an annotation model.
The annotation model(s) that have already been associated with an assembly will appear at the top of the Index to drop-down list. Choose the annotation model you wish to index to, select the Build index radio button and click Create (Figure 1). To build an aligner index based on an annotation model, a file must already be associated with the assembly.
Adding a Variant Annotation Database
Click the + Add variant annotations button under the Variant annotations section header on the .
If you are using a human - hg19 assembly, variant annotation databases from various sources will appear in the Variant annotation drop-down list (Figure 1). Available variant annotation database sources include:
dbSNP
Kaviar
Removing Library Files
This document will show you how to delete an assembly and how to dissociate a library file.
Task Management
Partek Flow supports multiple versions of analysis tools, including third-party software (e.g. aligners). By default, the most recent versions are implemented during installation of Partek Flow. In some circumstances, you may need to use a different version of a tool, e.g. if reproducing an older study. Furthermore, tasks can be enabled and disabled.
Task Management Page
Only Partek Flow users with administrator privileges have access to the Tasks management page. Changes made on this page are applied to all other users on the Partek Flow instance. To access the page, click the avatar in the top right corner, choose Settings and click Tasks on the left (Figure 1).
Figure 1. The drop-down list will show all assemblies that have a Bowtie 2 index associated with them. Choose New assembly... to add a Bowtie 2 index to another assembly
Figure 2. Add Bowtie 2 index to another assembly not listed in Figure 1.
Figure 3. If no gene sets are associated with the assembly, click New assembly... to add one.
Figure 4. Download a gene set database using the Create gene set database dialog from within a project
Figure 5. Import a gene set database using the Create gene set database dialog from within a project
Figure 3. When importing an annotation model, the data type must be specified
Figure 4. Select the column with gene- and transcript level information
Figure 5. Comparison of genome coordinate formats. The example column shows the coordinates for a feature that starts at position 1000 (inclusive) and stops at position 2000 (inclusive).
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Figure 1. Various versions of SnpEff variant databases are available for automatic download. The drop-down list shows databases available for human - hg38. Choose a SnpEff database from the drop-down list, select the Download database radio button and click Create.
Figure 1. For certain model organisms, automatic downloads of VEP databases are available
Figure 2. VEP databases can be added for other species
All pipelines that have been created or imported within the same Partek Flow instance can be shared by all users. The pipeline manager allows each user to choose the pipelines that appear on their Task Menu. To hide a pipeline, select the corresponding checkbox in the
Ignore
column (Figure 1).
Downloading a pipeline
To download a pipeline, click the Download pipeline in the Actions column. A file with the extension *.pipeline will be download to the default download directory of your local computer.
Deleting a pipeline
Pipelines can be deleted by the owner (as listed in the Creator column) or by a user with admin privileges. Clicking Delete pipeline in the Actions column will delete the pipeline and it will no longer be available to any Partek Flow user. Note that deleting a pipeline affects all users of the same Partek Flow instance.
Importing a Pipeline
Within the Pipeline management page, you can upload a pipeline from your local computer or import a pre-built pipeline directly from the Partek website. You can import a pipeline even before you create or open a project within Partek Flow.
From My Computer
To import a .pipeline file from your local machine, click on the + Import pipeline button on the Pipelines management page
By default, the My computer option at the top of the Import pipeline page should already be selected. Click the + Choose button (Figure 2)
Browse to the location of the .pipeline file on your local machine and select it for upload. The selected file name will be displayed next to the Choose pipeline button
If you wish to give the pipeline a new name, type a new name into the Rename pipeline box and click Import
Figure 2. Importing a pipeline from your local machine
From Partek Website
To download a pipeline from the Partek website, click on the + Import pipeline button
Select Hosted pipelines (Figure 3). This will display all available pre-built pipelines from the Partek website.
Click the Import pipeline button next to the pipeline you wish to import (Figure 3). Alternatively, click Import all pipelines at the bottom of the page to import all pipelines displayed.
Figure 3. Pre-built pipelines that can be imported from the Partek website. To preview a pipeline to see more detail, click the pipeline image
For additional information, visit the Pipelines documentation page.
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Figure 1. Pipeline Management Page view of an administrative user. The view for non-administrative users will not have the 'Enable by default' column and pipelines created by other users will appear as 'Anonymous'
If you are using an assembly supported by Partek (e.g. human), annotation models from a variety of commonly used sources will appear in the Index to drop-down list in addition to the ones that have already been associated with the assembly. If you choose an annotation model that has not already been associated, it will automatically be downloaded prior to building the index.
Figure 1. Building an aligner index for an annotation model
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Multiple versions of the above databases are available. For human - hg38, only dbSNP is currently available. This list is periodically updated.
Figure 1. For human - hg19, automatic downloads of various variant annotation databases are available from the Partek repository
Choose a database from the drop-down list, select the Download variant database radio button and click Create.
If you prefer to add a custom variant annotation database, perhaps from another source or 'gold-standard' validated variants, choose Add variant database from the Variant annotation drop-down list (Figure 2). Name the variant annotation database by typing into the Custom Name box and click Create. Characters such as $ * | \ : " < > ? / % cannot be used in custom names. A variant annotation database can be added from the Partek Flow Server or a URL download link. The behavior of each option is similar to when importing a reference sequence (see Adding a Reference Sequence). When browsing for files on the Partek Flow server, only the files with relevant file extensions will be visible (.vcf and various compressed formats).
Figure 2. Add a custom variant annotation database
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To delete an assembly from the Library File Management, select the Delete assembly () icon by the assembly name. You will be prompted to confirm the deletion (Figure 1). If you also want to delete the individual library files from the disk, select the Delete the library files check box. Select Delete to proceed with deletion. Note that the deletion can not be un-done.
Figure 1. Delete assembly dialog. To remove all the library files from the disk, select the Delete the library files option
Disassociate Library Files
Individual library files can be removed by disassociating them from their respective assembly. To disassociate a file, click on Disassociate library file in the Actions column (Figure 2).
Figure 2. Dissociate library file icon
If the library file is used by multiple projects, the resulting dialog will provide a list of projects (Figure 3). As long as the library file is used, the Delete the library files option will be disabled.
Figure 3. Dissociate library file dialog. The dialog lists all the projects using the selected library file
Using the Dissociate library file tool does not physically remove the file from the disk, but disassociates the file from the selected library file name. This, in turn, enables you to associate newer or improved version of the library file with the same library file name. If the goal is to physically delete the library files from the disk, then all the projects using that library file must be deleted first.
Figure 1. Task management page. The table shows one task per row and the current system wide version for each task
To enable or disable a task, select or deselect the checkbox next to the Task name on the Tasks management page, respectively (Figure 1).
To change the version of a task, click the drop-down and choose a different version from the drop-down list (Figure 2). All tasks launched after the change will use the chosen version.
Figure 2. Choose a new version from the drop-down list
To review which version has been used to perform a task, hover the cursor over the task node and inspect the pop-out balloon (Figure 3). Alternatively, the version number is shown on the Task details (Figure 4) page and the Data summary report (Figure 5).
Figure 3. Hovering over the task node shows the task version
Figure 4. The Task details page shows the task version (shown in bold)
Figure 5. The Data summary report shows the task version (shown in bold)
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Library Files
The library files associated with the selected assembly are organized into several sections. Below is some information on each section.
This section includes two types of library file: reference sequence and cytoband files.
Reference sequences are the chromosome/scaffold/contig DNA sequences for a species. A reference sequence file is typically in FASTA or 2bit format. The reference sequence of a species is used for aligner index creation, variant detection and visualization of the reference sequence in the Chromosome view.
Cytoband files are used for drawing ideograms of chromosomes in the , including positions of cytogenetic bands if known.
Reference aligner indexes
Next-generation sequencing aligners require the reference sequence to be indexed prior to alignment, as this greatly increases alignment speed. An index consists of a set of files (Figure 1) and are generally aligner specific. For example, if you wish to align using BWA, you need a BWA index.
Some of the supported aligners share indexes. If you want to align using Tophat, the Bowtie aligner indexes can be used. If you want to align using Tophat2, the Bowtie2 aligner indexes can be used.
Some aligner indexes are version specific, so care must be taken if you change aligner versions. For example, the index files for STAR version 2.4.1d are different to older versions of STAR.
This section contains aligner indexes for aligning to the whole genome. If you wish to align to a subset of the genome, e.g. targeted amplicons or the transcriptome, you must generate these indexes in the Annotation models section.
Gene sets
Gene set files are required for biological interpretation analyses (e.g. GO enrichment). Genes are grouped together according to their biological function. Gene set files have to be in GMT format, where each row represents one gene set. The first column of a GMT file is the GO ID or gene set name. The second column is an optional text description. Subsequent columns are the gene symbols that belong to each gene set. Gene ontologies for various model organisms are available for automatic download from the Partek repository (source: ). Because gene ontologies are frequently updated, is checked for updates quarterly. You can check for recent updates to the Partek repository .
Variant annotations
Variant annotation databases are collections of known genomic variants (e.g. single nucleotide polymorphisms). If you have performed a variant detection study, detected variants can be searched against variant annotation library files to see if the detected variants are known from previous studies. Furthermore, you can validate detected variants against 'gold-standard' variant annotation library files. Variant annotation files are typically in VCF format.
Variant annotation databases from commonly used sources (e.g. dbSNP) are available for automatic download from the Partek repository. Because variant annotation databases are frequently updated, these sources are checked for updates quarterly. You can check for recent updates to the Partek repository .
SnpEff variant databases
1 is a variant annotation and effect prediction tool that requires its own variant annotation files, separate to the other Variant annotation library files. If you wish to use SnpEff, library files need to be added to this section.
VEP database
The Ensembl Variant Effect Predictor () is another variant annotation and prediction tool that requires its own annotation files, separate to the Variant annotation library files. If you wish to use VEP, library files need to be added to this section.
Annotation models
This section includes two types of library file: annotation models & aligner indexes.
Annotation models describe genomic features (e.g. genes, transcripts, microRNAs) for a specific version of the reference sequence. Annotation models contain labels (e.g. gene ID) and genomic coordinates (e.g. chromosome, start & stop position) for each feature.
Annotation models will appear in separate tables (Figure 2). If you have multiple versions of annotation models from the same source, it is advisable to distinguish them by their date or version number.
Annotation models from commonly used sources (e.g. Refseq, ENSEMBL) are available for automatic download from the Partek repository. Because annotation models are frequently updated, these sources are checked for updates quarterly. You can check for recent updates to the Partek repository .
Annotation models are used for quantification in gene expression analyses, annotating detected variants (e.g. to predict amino acid changes), visualizations in , generating coverage reports and for aligner index creation (see ). Typical file formats include GTF, GFF, GFF3 and BED.
The arrows ( v /) next to the annotation model name expand/collapse each table. Two of the annotation models displayed in Figure 2 are different versions from the same source (Ensembl), distinguishable by their version number. Aligner indexes (e.g. for alignment to the transcriptome) are added to the table of the corresponding annotation model.
The aligner indexes in the Annotation models section are required if you wish to align to a subset of the genome as defined by the annotation model, e.g. target amplicons or the transcriptome. The reference sequence is still required to generate an aligner index for an annotation model. As with whole genome alignment, indexes are aligner specific, although some aligners share indexes and are version specific (see ). The aligner indexes generated will be added to the corresponding annotation model table (Figure 2).
References
Cingolani P. et al. A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso-2; iso-3. Fly. 6(2):80-92. PMID: 2272867
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Adding Prep kit
Partek distributes prep kit files for a variety of single cell technologies, such as 10x Genomics, Drop-seq, and Fluidigm C1. Prep kit files are required to process single cell fastq files. If you need to add a new prep kit in order to process your fastq files, you will need to get detailed information on how the library is constructed for the specific assay.
The following instructions use the 10X Genomics Chromium single cell gene expression 3' v3 chemistry as an example to illustrate how a prep kit tag library file is made in Partek Flow. Figure 1 is a schematic diagram of the single cell 3' v3.1 gene expression assay:
Click + Add prep kit button on Prep kit files tab at Library files management page, choose Other/Custom option from the Prep kit name drop-down list and specify a name e.g. Assay1 (Figure 2), choose the Build Prep kit option and click Create.
Figure 2. Add a new prep kit, name it Assay1
Select the Is paired end check button, and specify the information contained in each end:
Read 1: contains 10X barcodes and UMI, it is 28bp long. 10X Genomics has a which contains all known barcode sequences during library preparation. You will need to make a .fasta file format of this information and click on the + icon in the First mate segmentation (Figure 3) to add the barcode as the first segment in read 1.
Specify the new segment (Figure 4), choose Barcode from the Type drop-down list, specify the barcode .fasta file. If the barcodes are random with a fixed length, choose Arbitrary and specify the length. You can also choose the Manual option to manually add the barcode sequences that are available for this assay. Specify number of mismatches are allowed. Click Add
Click on the remove (-) icon to remove a segment if you make a mistake (Figure 5). Click on the + icon next to Barcode to add more segments, e.g. UMI.
According to the 10X 3' v3 assay, the UMI is a 12bp sequence (Figure 6).
Read 2 contains the 90bp insert sequence, click on the + icon next to Second mate segmentation, choose Insert from the Type drop-down list and specify the minimum read length allowed (Figure 7).
Check the Remove poly-A tail button and click Finish. The new prep kit file is added to the library file database in Partek Flow (Figure 8).
Click the three dots under Action, select View prep kit details to view the diagram (Figure 9).
When you perform the trim tag task on fastq files, Partek Flow will remove read 1 and the downstream alignment task will only be performed on read 2. The barcode and UMI information will be added in the read name in the output file for downstream analysis.
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Adding Library Files on the Library File Management Page
Library files can be added to an assembly by clicking the + Add reference files button under the Reference files section (Figure 1) on the library file management page. If all possible library files are already associated in the section, the Add reference files button will be disabled (Figure 1).
Figure 1. The Add reference files button is used to add library files to a section on the library file management page (top). The button is disabled if all possible library files have been added to a section (bottom).
Clicking + Add reference files button opens a dialog specific to the section. Choose a library type from the drop-down list in the Add reference sequence dialog (Figure 2) and the dialog will present options specific for each type. If certain library files have already been associated with an assembly, they will not appear as options in the dialog. For example, if a reference sequence has already been associated with an assembly, it will not appear as an option in the drop-down list shown in Figure 2.
Figure 2. Add reference sequence dialog
All library file tasks are logged in the System queue, which can be accessed by clicking on the avatar in the top right corner, choosing Settings and clicking System queue on the left. When a library file is being created, the hourglass () icon will appear in the Library file column. At the Action column, selecting View running tasks details will display information on the task progress (Figure 3).
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Lists
Create, view, and edit lists within a Partek Flow instance. To access Lists:
Option Set Management
Partek Flow allows you to configure advanced settings for various tasks including adapter trimming (Cutadapt), alignment (various aligners), quantification (Cufflinks), differential gene expression analysis (GSA, ANOVA and Cuffdiff), variant detection (Partek, samtools and Freebayes) and variant annotation (SNPEff). User-defined option sets can be saved and applied to other projects.
Click the user avatar in the top right corner of the Partek Flow interface
Choose Settings in the menu
Select Lists from the left panel of the Components section.
Lists can be accessed by administrative and non-administrative users (Figure 1).
Figure 1. Access Lists in the Partek Flow interface
View and sort the name of the list, number of features, owner, and creation date by clicking the column title of the list table. Each list has the option to ignore and actions to download list, edit the list name or description, share list or delete the list.
Create a new list
To create new list, select the + New list button on the top of the page and give the list a name. Choose between the three list options:
Local file: text file (.txt) on the local computer
Text: manually specify a list of names
Hosted lists: lists hosted on Partek website
When manually specifying a local file or text, only the user who added the list can view and use the lists. Hosted lists can be viewed and used by all of the users.
The list name needs to be unique for each user. A description is optional. The list of features can be specified from a .txt file on your local computer by selecting the Local file option (Figure 2). The .txt file should contain only one feature per row.
Figure 2. Adding a list from a .txt file
To manually enter a list of genes, select the Text option (Figure 3). Content can be typed into the text field or pasted from the clipboard as one feature per row.
Figure 3. Type or paste feature names in the text field in column format
Select Add list to add the list. The new list will be added to the list table.
When the Hosted lists option is chosen, lists can be downloaded from Partek (Figure 4).
Figure 4. Download a list from Partek
Click the check box for each list on the left to select lists and click Add list. The downloaded lists are automatically shared with all users on the server. If you do not want to see the list when performing tasks related to lists (e.g. filter feature task), check the Ignore button.
Once a list is added to the Partek Flow server, it can be used to filter data (e.g. to filter quantified RNA-Seq data to only include or exclude the genes in a list) or to color by a feature list.
Use a Feature list to Select & Filter populations
Feature lists can be used to Select & Filter.
In the Data Viewer, select the Style configuration option
Choose Feature list from Color by drop-down list
Select the List of interest using the drop-down
Choose a Metric for the feature list
Click Select & Filter under Tools on left
Use the Add criteria drop-down to choose Gene score
Use the slider or text fields to select a population
Use Filter at the top right to include () or exclude () this population
Apply observation filter can then be used to filter this population to a specific node in the analysis pipeline
To remove this filter, click Clear filters.
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A Bowtie alignment task will be used as an example, but the steps are the same for any task that allows configuration of advanced settings. Click Configure in the Advanced options section on the Task setup page (Figure 1).
Figure 1. Access to advanced options page through the Configure link. The example shown is on a Task setup page of a Bowtie alignment
In the task-specific Advanced options dialog, hover the cursor over the tool tip () icon to get more information on each setting. Make the desired changes to the settings and click Save as new (Figure 2). Note that clicking Apply will implement the custom option set for the current task, but the option set will not be saved for use in other projects.
Figure 2. Advanced options for Bowtie alignment task. The tool tip describes the setting in a pop-out balloon. In this example, the seed length has been changed from the default 28 to 10
Name the new custom option set, add a description (optional) and click Save in the New option set dialog (Figure 3).
Figure 3. Name and describe the new custom option set
The saved option set will appear in the drop-down list in the Advanced options section on the Task setup page (Figure 4). All saved option sets are available to all of the users of the same Partek Flow instance.
Figure 4. To apply a saved option set, choose it from the drop-down list in the Advanced options section of the Task setup page
Option Set Management Page
To access the Option set management page, click the avatar in the top right corner, choose Settings and click Option sets under Components section on the left (Figure 5). This page can be accessed by administrative and non administrative Partek Flow users.
Choose a task from the selector and all associated option sets will be shown in a table (Figure 5). Click the three dots in the Actions column and select View parameters to display the option set (Figure 6). Select Delete option set in the Actions column to delete an option set. Note that only the Partek Flow user who created an option set and administrative Partek Flow users are able to delete an option set.
Figure 5. Option set management page displaying a Bowtie task option set
Figure 6. View an option set
Adding an Option Set from the Command Line
This is a Partek Flow licensed feature separate from the base license.
User-defined command lines can be copied and pasted to create a custom option set. This is particularly useful if, for example, you have previously run an alignment with custom options on the command line and wish to use the same set of options for an alignment in Partek Flow.
On the Option set management page, click the + Add option set from command line button (Figure 5). In the Add option set dialog (Figure 7), choose a task from the Select Task drop-down list and the version number (if applicable). Name the option set by typing into the Option set name box. Add a description of the option set by typing into the Option set description box (optional). Paste the command line from the clipboard into the Command line box and click Save.
Figure 7. Add option set dialog
The command line will be parsed, validated and converted into a Partek Flow option set. The Command line parsing report will summarize the Command, Recognized parameters and Ignored parameters (Figure 8). Partek Flow will ignore parameters with a syntax error or parameters already dealt with by other components of the software (e.g. input/output file paths, performance optimization). Click OK to close the report. The option set will now appear on the Option set management page (Figure 5) and can be applied to projects (Figure 4).
Figure 8. Command line parsing report
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Partek Flow enables certain components of the software to be configured, such as hiding/showing special tasks and managing saved task parameters like option sets.
This section will illustrate how to use the following options:
Note that this task is for adding indexes for alignment to the whole genome. If you want to align to the transcriptome or another set of genomic features, see Adding Aligner Indexes Based on an Annotation Model.
Click the + Add reference aligner indexes button under the Reference aligner indexes section header on the library file management page. Choose the aligner index you wish to add from the drop-down list in the Add aligner index dialog (Figure 1). The following indexes can be added:
Bowtie
Bowtie 2
HISAT 2
TMAP
BWA
GSNAP
GSNAP v8
STAR
STAR 2.5.3a
STAR 2.7.3a
STAR 2.7.8a
Issac 2
RSEM
Salmon
Cell Ranger ARC 2.0.0 reference
Minimap2
If you are using an assembly supported by Partek (e.g. human), there are three radio button options: Download index; Build index or Import index (Figure 1). Certain aligner indexes may not be available for automatic download because the file sizes are too large to download efficiently.
If available, select Download index and click Create to get the chosen reference aligner index from the Partek repository.
Alternatively, select Build index and click Create to build the reference aligner index. To build an aligner index, a reference sequence file must already be associated with the assembly. Depending on the aligner, you may have to specify further parameters. Consult the user documentation for each aligner for guidance (usually available on-line).
Alternatively, select Import index and click Create to add an aligner index from another source. An aligner index can be added from the Partek Flow Server or a URL download link. The behavior of each option is similar to when importing a reference sequence (see ). When browsing for files on the Partek Flow server, only the files with relevant file extensions will be visible. This will vary for each aligner.
For custom assemblies (e.g. for non-model organisms), only the Build index and Import index options are available (Figure 2).
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