Differential Expression and Pathway Analysis
The Differential Expression and Pathway Analysis (
) and the Differential Expression and Pathway Analysis from Count Matrix (
) template workflows can be used for performing differential expression analysis for gene or transcript expression data from groups of samples.
The two workflows include all the necessary steps for the analysis:
- Differentially expressed features (genes or transcripts) are identified using Differential Expression for RNA-Seq, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Differential_Expression_RNA_Seq.html.
- The differentially expressed features are optionally uploaded to QIAGEN Ingenuity Pathway Analysis (IPA) using Upload to IPA.
- Terms from a Gene Ontology Annotation (GOA) table affected by the differentially expressed features are identified using Gene Set Test, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Gene_Set_Test.html.
- The expression values are summarized in various plots using
- PCA for RNA-Seq, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=PCA_RNA_Seq.html;
- Create Sample Level Heat Map for RNA-Seq, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Create_Sample_Level_Heat_Map_RNA_Seq.html;
- Create Feature Level Heat Map for RNA-Seq, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Create_Feature_Level_Heat_Map_RNA_Seq.html.
The two workflows differ in the accepted inputs:
- Differential Expression and Pathway Analysis takes as input Gene Expression (GE) or Transcript Expression (TE) tracks. These must be associated with a CLC Metadata Table, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Metadata.html for details.
- Differential Expression and Pathway Analysis from Count Matrix imports Gene Expression (GE) or Transcript Expression (TE) tracks, along with a CLC Metadata Table, from two separate files containing the expression data and metadata, respectively. See https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Import_Expression_Data.html for details.
Launching the workflows
To run these workflows, go to:
Template Workflows | Biomedical Workflows (
) | Whole Transcriptome Sequencing (
)
and select:
Differential Expression and Pathway Analysis ()
Differential Expression and Pathway Analysis from Count Matrix ()
For general information about launching workflows, see https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Launching_workflows_individually_in_batches.html
The input options can be configured in the following dialogs:
- For the Differential Expression and Pathway Analysis from Count Matrix workflow:
- Select Expression Tracks (TE or GE).
Select Gene Expression (GE) or Transcript Expression (TE) tracks.
- Configure batching. If running the workflow in Batch mode, you will be asked to define the batch units. See https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Running_workflows_in_batch_mode.html for details.
- Select Expression Tracks (TE or GE).
Select Gene Expression (GE) or Transcript Expression (TE) tracks.
- For the Differential Expression and Pathway Analysis from Count Matrix workflow:
- Import Expression Data. Select the two files containing the expression data and metadata, respectively.
Options in the following dialogs can additionally be configured for both workflows:
- Choose where to run. If you are connected to a CLC Server via the CLC Workbench, you will be asked where you would like to run the analysis. We recommend that you run the analysis on a CLC Server when possible.
- Specify workflow path. Select whether you want to upload the differential expression data to IPA. Uploading requires an IPA account.
- Specify reference data handling.
Select the relevant Reference Data Set, see Reference data management for details.
- Differential Expression for RNA-Seq.
Configure the following options as needed:
- Metadata table
- Test differential expression due to
- While controlling for
- Comparisons
- Control group
- If uploading to IPA:
- Filter on Custom Criteria. Configure filtering criteria defining which features to upload to IPA. See https://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Filter_on_Custom_Criteria.html for details.
- Upload to IPA. Login to IPA and configure the options. See Upload to IPA for details.
- Result handling. Choose if a workflow result metadata and/or log should be saved.
- Save location for new elements. Choose where to save the data, and press Finish to start the analysis.
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