![]() ![]() Under "Basic fields" -> Enrichment statistics-> select "weighted" *.Gene sets database: open-> click "Gene matrix(local gmx/gmt)" -> select "bp.gmt" -> "OK".Use FileZilla to download the two data files bp.gmt & genes.rnk from the directory /shared_data/RNAseq-function/.Ĭlick "Load data", then click "Browse for files", and open the two files bp.gmt & genes.rnkĬlick "Run GSEA Preranked", and set the following: You can run GSEA either on your laptop, or on the LINUX server. You can get the data from DEseq2, removing genes with expression level too low (filter by the DEseq2 output column baseMean) The order of genes in this file does NOT matter, as GSEA will sort this file based on column 2. ![]() Briefly, it is a tab delimited text file with two columns: gene name & log2(FoldChange). rnk: The file format is explained in the same web site as "".gmt". If you start with a GO annotation file like the one downloaded from Ensembl, use this procedure to convert to. When you work on your own project, here is how to prepare the two files. We provide you with two data files (bp.gmt & genes.rnk) to run GSEA. When you write "method" section of your manuscript, you can write that "gene set enrichment analysis was performed with BioConductor topGO package, using its default weight01 algorithm and topgo Fisher test, and p-value threshold is set at. The enriched GO ids are sorted by p-values in the column "topgoFisher". You can copy-paste each section in this file into an Excel spreadsheet (using text import wizard with "fixed width"). The myBP file can be opened with a text editor. You can use FileZilla to download both files to your laptop to examine the contents. myBP_Topgo_weight01_134_all.pdf: a PDF file with tables and plots.You can also test for MF (molecular function) and CC (cellular component).Īfter this, you should find two new files "myBP". TopgoAnnot, refset and DElist_pos: three input files Ġ.1: cutoff p-value for reporting enriched GO categories.īP: test for biological processing GO. programs/R-4.0.0/bin/Rscript /shared_data/RNAseq-function/topGO.r topgoAnnot refset DElist_pos 0.1 BP myBP We provide you with a script that converts the BioMart file into topGO format. Use FileZilla to upload the "mart_export.txt" file to your assigned Linux server, keep it in the directory "/workdir/$USER/exercise3". You will be prompt to save the file as "mart_export.txt".Ģ.2 Convert the GO annotation file into a format compatible with topGO. At the top of the page, click "Results".In the right panel, expand "EXTERNAL", check " GO term accession".In the right panel, expand "GENE", make sure "Gene stable ID" is checked, and.From the pull-down menu "CHOOSE DATASET" select "Sacchromyces cerevisiae".From the pull-down menu "CHOOSE DATABASE" select "Ensembl Genes xxx".Use a web browser to navigate to Ensembl web site:, and click BioMart.You will perform ORA on RNA-seq results of a yeast experiment.Ģ.1 Download GO annotation from Ensembl BioMart.Įnsembl ( ) is a good resource to retrieve existing GO annotation (For plant genomes, go to ) The topGO package of Bioconductor will be used to do ORA. Search for "EnrichmentMap" and install this app. After installation, you will need to install an app within Cytoscape named "EnrichmentMap". Go to and click "Download x.x.x" to download and install the software. Go to and click "Downloads" to download and install the software. ![]()
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