Analyze Count Matrix and Upload Comparisons to IPA
The workflow Analyze Count Matrix and Upload Comparisons to IPA imports expression data from an Expression count matrix, and analyzes them using the tools in the RNA-Seq Analysis folder. It then submits the comparison to IPA using the Pathway Analysis tool.
The purpose of the workflow is to make it as easy as possible for the user to get from Sample to Insight. The user only has to provide the expression count matrix, sample metadata as well as mRNA and Genes as input, and the workflow performs import, statistical analyses and data interpretation using capabilities available via CLC Genomics Workbench and IPA.
Opened in the workflow editor, the workflows looks like this (see figure 4.2 below):
Figure 4.2: Layout of the Analyze Count Matrix and Upload Comparisons to IPA workflow.
The expression data from the count matrix data are imported by the Import Expression Data tool that splits each sample into a track. The expression tracks are then sent to three tools:
- Create Heat Map for RNA-Seq The tool creates a two dimensional heat map of expression values. Each column corresponds to one sample, and each row to a feature (a gene or a transcript). The samples and features are both hierarchically clustered.
- Differential Expression for RNA-Seq The tool performs a statistical differential expression test for a set of Expression Tracks. It's outputs are used as inputs for the IPA tool and for Create Venn Diagram for RNA-Seq (see below).
- PCA for RNA-Seq The tool creates a PCA plot, which is a projection of a high-dimensional dataset (where the number of dimensions equals the number of genes or transcripts) onto two of three dimensions. This helps in identifying outlying samples for quality control, and gives a feeling for the principal causes of variation in a dataset.
The outputs from the tools are saved in the chosen output folder for the workflow. The outputs from the Differential Expression for RNA-Seq tool are furthermore used for processing by these two tools:
- Pathway Analysis The tool uploads the comparisons to IPA. See Uploading data to IPA using the Pathway Analysis tool for details.
- Create Venn Diagram for RNA-Seq The tool makes it possible to compare two or more statistical comparison tracks. The Venn diagram comparison visualizes the overlap between the differentially expressed genes or transcripts in the selected statistical comparison tracks. The genes considered to be differentially expressed can be controlled by setting appropriate p-value and fold change thresholds.
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