Tutorial 2.3. Identifying significantly upregulated genes

Run Oncodrive to identify the genes that are significantly up-regulated taking into account a set of tumour samples

Files needed

• gse19188_l2r-cancer-vs-normal.cdm.gz: which contains log2 ratios of tumour samples versus normal controls.

Run Oncodrive analysis

• See this chapter for details on how to run an Oncodrive analysis
• Select gse19188_l2r-cancer-vs-normal.cdm.gz as data file
• In filtering options choose “Transform to 1 (0 otherwise) cells with values greater than 1.297 (this is the optimal cutoff for this dataset obtained as explained in the supplementary material of IntOGen paper).
• Do not select anything in the sets of columns window.
• Leave the default values in the statistical test window (binomial, and Benjamin Hochberg FDR).
• Give a name to the analysis. Select a directory where to save it and click Finish.
• In the analysis details tab, click on "heatmap" under "Results" to view the heatmap of the results.

Import annotations for probes using Gitools

• Import Ensembl gene ids, symbol and description for probes in the Affymetrix HG U133 Plus 2 platform.
• Go to the Welcome tab and click the Biomart wizard. Choose "Table" and click Next.
• Select "Ensembl Genes 62" and "Homo Sapiens", and then the desired attributes. Leave only "Ensembl Gene ID" from the defaults, and add "Description" (under Gene/ENSEMBL), "Associated gene name" (under Gene/ENSEMBL) and "Affy HG U133-PLUS-2" (under microarray).
• Bring up in the list the Affymetrix attribute and click Next.
• Do not apply any filters, and skip rows with empty values.
• Give a meaningful name to the file, select an output folder where to save it and finish.
• Load the annotations for rows by going to the rows tab under properties and loading the annotation file. Click "Filter".
• Add a header in the rows tab and select "Text labels", then "Ensembl Gene ID" and finish. Do the same for the other two annotations.
• Bring up the header for "Associated Gene name" and sort by this label.
  
• Alternatively, you can download the Affymetrix HG U133 Plus 2 ids from here:
  • homo_sapiens_affy_u133_plus_2_annotations.tsv

Explore the results