SigTerms tutorial: using the “FindSignificantTerms” macro Chad Creighton, Ph.D. Selecting the Annotation workbook • The SigTerms web site provides links to download pre-compiled Annotation workbooks for several types of gene class associations of potential interest (e.g. GO terms, microRNA targeting predictions). • Annotation workbook should be specific to the gene array that was used for the profiling. – All genes in the workbook should be represented on the array (any genes not on the array should not be represented in the Annotation workbook) – Workbooks specific to particular array platforms are available for download • User can construct his or her own Annotation workbook and specify gene-to-term associations – Details on Annotation workbook format on web site. • Open both SigTerms.xls and Annotation workbook in Excel • Make sure your Excel allows the running of macros (see web site for details). • In the Annotation workbook, insert a new worksheet • (Right-click on the worksheet tab, select “Insert”, and choose “New Worksheet.) • In the new worksheet, paste the list of selected genes in the first column (starting from the first row). • Use Entrez gene number to specify gene – Duplicate Ids are okay. – For statistical calculations, should not include NULL or “---” or “///” containing entries, only entries for genes with Entrez numbers. • Run the “FindSignificantTerms” macro. • Selected gene worksheet should be “active” (i.e. at the front of all the other sheets) when running the macro. • Alt+F8 or Tools->Macros (32-bit Excel) displays the SigTerms macros. • Specify the gene population – If choose “number of entries in Annotation sheet”, then selected gene list should not have any entries not in the Annotation worksheet – Otherwise, specify the number of unique, identified genes on the array (for pre-compiled Annotation workbooks, “Gene Pop” sheet has this number). • “Enriched Terms” output worksheet lists gene class terms ranked by statistical enrichment • “Terms with Genes” output worksheet lists, for each term, the genes that fall under that term. • If desired, one can link additional information to the genes (or microRNAs) listed in the output worksheets. • Can use “MATCH” and “INDEX” Excel functions (Illustrated above and in our Excel tutorial).