1. Getting GO: how to get GO for functional modeling Iowa State Workshop 11 June 2009

2. 1. GO for your species  GOProfiler: summarizing the available GO 2. GO browsers  QuickGO from EBI  AmiGO from the GO Consortium 3. gene association files 4. getting GO for your dataset 5. adding more GO 6. requesting GO 7. GO based tools for functional modeling

3. 1. GO for your species

4. GOProfiler GOProfiler allows you get an overview of what GO annotation exists for the species you are interested in.

8. Number of proteins is based upon UniProtKB records for these species. Species with only IEA annotations do not have an active GO annotation project  GO provided automatically by EBI GOA Project.

9. 2. GO Browsers

10. Use GO Browsers for: searching for GO terms searching for gene product annotation filtering sets of annotations and downloading results creating/using GO slims

11. GO Browsers QuickGO Browser (EBI GOA Project)  http://www.ebi.ac.uk/ego/ http://www.ebi.ac.uk/ego/  Can search by GO Term or by UniProt ID  Includes IEA annotations AmiGO Browser (GO Consortium Project)  http://amigo.geneontology.org/cgi- bin/amigo/go.cgi http://amigo.geneontology.org/cgi- bin/amigo/go.cgi  Can search by GO Term or by UniProt ID  Does not include IEA annotations More information about these tools is available from the online workshop resources.

12. 3. gene association files

13. The gene association (ga) file standard file format used to capture GO annotation data tab-delimited file containing 15* fields of information:  Information about the gene product (database, accession, name, symbol, synonyms, species) information about the function:  GO ID, ontology, reference, evidence, qualifiers, context (with/from) data about the functional annotation  date, annotator * 2 additional fields will soon be added to capture information about isoforms and other ontologies.

15. (additional column added to this example)

16. gene product information

17. metadata: when & who

18. function information

22. Gene association files GO Consortium ga files  many organism specific files  also includes EBI GOA files EBI GOA ga files  UniProt file contains GO annotation for all species represented in UniProtKB AgBase ga files  organism specific files  AgBase GOC file – submitted to GO Consortium & EBI GOA  AgBase Community file – GO annotations not yet submitted or not supported  all files are quality checked

23. 4. Finding GO for your dataset

24. The AgBase GO annotation tools can be used separately or can be combined to rapidly provide an annotation file for functional modeling tools.

26. GORetriever Allows you to get GO annotations for a specific set of gene products. Accepts a text file of UniProt accessions or IDs or gi numbers. Returns GO annotations, list of accessions that had no GO and a GO Summary file.

27. GORetriever Results

29. save as text file For GOSlimViewer

30. GORetriever Results

31. But what about IDs not supported by GORetriever?

32. 5. Adding GO to your dataset

33. only returns existing GO only accepts limited accession types GOanna does a Blast search against existing GO annotated products. allows you to quickly transfer GO to gene products where they have similar sequences (ISS) accepts fasta files

34. GOanna

37. GOanna Results

38. query IDs are hyperlinked to BLAST data (files must be in the same directory)

39. *WHAT IS A GOOD ALIGNMENT? 1. Manually inspect alignments and delete any lines where there is not a good alignment*. 2. Add this additional annotation to the annotations from GORetriever.

40. GOanna2ga New to AgBase: an online script to convert your GOanna file to a gene association file format. Allows you to add manually checked GOanna annotations to a GORetriever file. Link is available from the workshop resources.

41. 6. Requesting GO

44. 7. GO based tools for biological modeling

45. GOSlimViewer: summarizing results

46. response to stimulus amino acid and derivative metabolic process transport behavior cell differentiation metabolic process regulation of biological process cell communication nucleobase, nucleoside, nucleotide and nucleic acid metabolic process cell death cell motility macromolecule metabolic process multicellular organismal development catabolic process biological_process “process unknown” “function unknown” “component unknown” ??

47. http://www.geneontology.org/

49. However…. many of these tools do not support agricultural species the tools have different computing requirements Tools for GO analysis of gene expression/microarray data A list of these tools that can be used for agricultural species is available on the workshop website at the Expression analysis tools at the GO consortium website link.

50. Evaluating GO tools Some criteria for evaluating GO Tools: 1. Does it include my species of interest (or do I have to “humanize” my list)? 2. What does it require to set up (computer usage/online) 3. What was the source for the GO (primary or secondary) and when was it last updated? 4. Does it report the GO evidence codes (and is IEA included)? 5. Does it report which of my gene products has no GO? 6. Does it report both over/under represented GO groups and how does it evaluate this? 7. Does it allow me to add my own GO annotations? 8. Does it represent my results in a way that facilitates discovery?