Functional And Pathway Analysis 2010

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The Functional and Pathway Analysis talk given in March 2010 at the CRUK CRI. Cambridge UK.

It was designed to introduce wet-lab researchers to using web-based tools for doing functional analysis of gene lists, such as from microarray experiments.

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Functional And Pathway Analysis 2010

  1. 1. Functional and Pathway Analysis Stewart MacArthur Bioinformatics Core March 18th, 2010 Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 1 / 19
  2. 2. Introduction The Problem The Problem • High-throughput genomics methods: • microarrays • next generation sequencing • Generate large lists of “interesting” genes • How to we summarize? • What are the themes of the lists? Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 2 / 19
  3. 3. Introduction The Solution The Solution • Functional Analysis • Determine common functions • Find groups of functionally related genes • Pathways Analysis • Determine common pathways • Determine potential up/down stream regulators Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 3 / 19
  4. 4. Enrichment Analysis Methods The Methods Enrichment Analysis Are there more of the genes in my list in functional category X than we could expect by chance? • SEA - Singular Enrichment Analysis • MEA - Modular Enrichment Analysis • GSEA - Gene Set Enrichment Analysis Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 4 / 19
  5. 5. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  6. 6. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  7. 7. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  8. 8. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  9. 9. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  10. 10. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  11. 11. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  12. 12. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  13. 13. Enrichment Analysis Methods Hypergeometric Brief Aside: Hypergeometric Test The hypergeometric test calculates the probability that the number of genes in our gene list that are in functional category/pathway X occured by chance Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 5 / 19
  14. 14. Enrichment Analysis Methods SEA - Singular Enrichment Analysis SEA - Singular Enrichment Analysis Inputs: • List of “interesting” genes, e.g. DE genes • List of functional annotations e.g. GO annotations Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 6 / 19
  15. 15. Enrichment Analysis Methods SEA - Singular Enrichment Analysis SEA - Singular Enrichment Analysis Inputs: • List of “interesting” genes, e.g. DE genes • List of functional annotations e.g. GO annotations Method: For each annotation • Are more of the genes in our list present than would be expected by chance • Calculate p-value Next annotation • Correction for multiple testing Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 6 / 19
  16. 16. Enrichment Analysis Methods SEA - Singular Enrichment Analysis SEA - Singular Enrichment Analysis Inputs: • List of “interesting” genes, e.g. DE genes • List of functional annotations e.g. GO annotations Method: For each annotation • Are more of the genes in our list present than would be expected by chance • Calculate p-value Next annotation • Correction for multiple testing Output: • Ranked list of annotations Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 6 / 19
  17. 17. Enrichment Analysis Methods MEA - Modular Enrichment Analysis MEA - Modular Enrichment Analysis • Extension of SEA • Incorporates network discovery algorithms • Considers term-to-term relationships • Terms not treated as separate tests • Uses co-occurrences of terms • More closely related to biology • Based on assumption that related functional groups have similar member genes Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 7 / 19
  18. 18. Enrichment Analysis Methods GSEA -Gene Set Enrichment Analysis GSEA No cutoff, uses all genes ranked e.g. microarray experiment ranked by fold change or differential expression For each functional annotation • Are genes randomly distributed in ranked list? or • Are genes distributed towards the top/bottom of the list? • Calculate enrichment score (ES) • Calculate significance of ES Next annotation • Correct for multiple testing Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 8 / 19
  19. 19. Enrichment Analysis Methods GSEA -Gene Set Enrichment Analysis GSEA Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 9 / 19
  20. 20. Enrichment Analysis Methods GSEA -Gene Set Enrichment Analysis GSEA Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 9 / 19
  21. 21. Enrichment Analysis Methods GSEA -Gene Set Enrichment Analysis GSEA Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 9 / 19
  22. 22. Enrichment Analysis Methods GSEA -Gene Set Enrichment Analysis GSEA Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 9 / 19
  23. 23. Annotation Resources Annotation Resources Where do the gene sets come from? • GO - Gene Ontology • KEGG - Kyoto Encyclopedia of Genes and Genomes • MSigDB - Molecular Signatures Database • Pathway Commons • ... • ... Choice of annotation often dictated by choice of tool Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 10 / 19
  24. 24. Web based tools Tools • Approximately 68 enrichment tools Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 11 / 19
  25. 25. Web based tools Tools • Here they are: Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 11 / 19
  26. 26. Web based tools Tools • Mainly Web based Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 11 / 19
  27. 27. Web based tools Tools • Mainly Hypergeometric based Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 11 / 19
  28. 28. Web based tools Recommended Tools • SEA - ClueGO, GOStat, • MEA - DAVID, GOToolBox • GSEA - GeneTrail, FatiScan (Babelomics) See Bioinformatics Core Wiki Page for more tools http://criwiki.cancerresearchuk.org/ Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 12 / 19
  29. 29. Web based tools David DAVID http://david.abcc.ncifcrf.gov The Database for Annotation, Visualization and Integrated Discovery • Over 1,600 DAVID citations • 37 nature-branded citations to date • Daily Usage: 1200 gene lists/sublists • Daily Usage: 400 unique researchers. Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 13 / 19
  30. 30. Web based tools David DAVID http://david.abcc.ncifcrf.gov The Database for Annotation, Visualization and Integrated Discovery • Identify enriched biological themes • Discover enriched functional-related gene groups • Cluster redundant annotation terms • Visualize genes on BioCarta & KEGG pathway maps • Search for other functionally related genes not in the list • Convert gene identifiers from one type to another. • And more Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 14 / 19
  31. 31. Web based tools GeneTrail GeneTrail Annotations include • KEGG • TRANSPATH • TRANSFAC • GO Methods: • Over-Representation Analysis (ORA) • Gene Set Enrichment Analysis (GSEA) Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 15 / 19
  32. 32. Commercial Tools Ingenuity Pathways Analysis (IPA) Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 16 / 19
  33. 33. Commercial Tools GeneGo MetaCore Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 17 / 19
  34. 34. Commercial Tools Suraj - GeneGO Demo Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 18 / 19
  35. 35. Cytoscape Cytoscape Stewart MacArthur (Bioinformatics Core) Functional and Pathway Analysis March 18th, 2010 19 / 19

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