Advancing Science with DNA Sequence Data Curation in IMG-ER Natalia Ivanova MGM Workshop May 16, 2012

Advancing Science with DNA Sequence Tricky question What do you need to do data curation in IMG? a)I-phone b)PhD in Computer Science c)supernatural powers Correct answer: you need an IMG account

Advancing Science with DNA Sequence 1.Gene models a)Add a gene b)Make a gene pseudogene or “obsolete” (=delete it) 2. Functional annotations: a)Product names b)EC numbers c)Gene symbols If you believe something else needs to be changed (genome name, taxonomy, etc.) – please use IMG Questions/Comments link What can’t be changed: automated assignments to protein families (Pfam, COGs, TIGRfam, InterPro, SEED assignments, KO assignments) What can be curated in IMG-ER?

Advancing Science with DNA Sequence Center point for curation – Gene Cart

Advancing Science with DNA Sequence Product Name is free text (but see GenBank requirementsProduct Name is free text (but see GenBank requirements mesubmit_annotation.html ) Prot Description is free text (goes to “note” in GenBank submission) EC number and PUBMED ID – see explanation Notes are free text (goes to “note” in GenBank submission) Gene symbol is “gene name” – 4 letter abbreviation; goes to “gene” in GenBank submission

Advancing Science with DNA Sequence How to find the genes that need curation? Two possible scenarios: You have submitted a genome to IMG-ER and want to have the best annotations possible for it (e. g. for GenBank submission)You have submitted a genome to IMG-ER and want to have the best annotations possible for it (e. g. for GenBank submission) You’re an expert and know everything about a certain pathway or protein family (families) = “community service”You’re an expert and know everything about a certain pathway or protein family (families) = “community service”

Advancing Science with DNA Sequence Curation of genome annotations Compare Gene Annotations find genome Genome Statistics review Gene Pages add to Gene Cart refine gene set Find Genomes: Genome Browser Genome Search “Hypothetical protein”, but with some evidence Non-hypothetical protein, but no evidence w/o enzymes but with candidate KO based enzymes Protein families Homologs/orthologs Gene Neighborhoods

Advancing Science with DNA Sequence Why do you want to review annotations? Most IMG pipelines are optimized for specificity, so they are more likely to have false negatives, but generate few false positives Compare Annotations –Product name is a consensus of multiple assignments: BLASTp, TIGRfam, COG, Pfam –Sources of false negatives - cutoffs: TIGRfam trusted cutoffs are quite stringent; COG doesn’t have trusted cutoffs; BLASTp cutoff of 50% identity Candidate genes with KO annotations – sources of false negatives –Cutoffs for % identity and alignment length

Advancing Science with DNA Sequence Curation of annotation in one genome (or a set of genomes) a)Your favorite genes (experimental verification, etc.) -> use Find Genes, Gene Search or BLAST b)“Compare Annotations” on Organism Details page c)“Candidate genes with KO annotations” on Organism Details page d)KEGG Pathways (either from Organism Details page or from Find Functions menu) e)PhyloProfiler

Advancing Science with DNA Sequence A shortcut for product name/EC number assignments based on KO

Advancing Science with DNA Sequence Example of a missed gene Run PhyloProfiler of Deinococcus geothermalis as a query, Deinococcus hopiensis as target (with no homologs in) Select Dgeo_0119 as a sequence to check whether a homolog of this gene was missed in Deinococcus hopiensis

Advancing Science with DNA Sequence Adding missed genes - contd Use graphical viewer to check the translation Adjust the start if other start codons with better RBS exist upstream

Advancing Science with DNA Sequence Reviewing your annotations Organism Details page -> Genome Statistics MyIMG

Advancing Science with DNA Sequence IMG curation exercises Go to the link in the usual place: The first 2 pages – questions without answers; the rest is cheat sheet