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Utilizing NCBI Assets for Quality Revelation

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  1. Using NCBI Resources for Gene Discovery Kim D. Pruitt Transcriptome 2002 National Center for Biotechnology Information (NCBI) National Library of Medicine National Institutes of Health http://www.ncbi.nlm.nih.gov/

  2. Fundamental resources Primary Databases - GenBank, dbEST, dbSTS, PubMed • Archival - original data submissions • Database staff organize, but don’t add additional information Derivative Databases – RefSeq, LocusLink, UniGene, Map Viewer • Curated/expert review • compilation and correction of data • Computationally Derived • Combinations

  3. Infrastructure Sequences Structure Integrating Information To Facilitate Discovery, Retrieval, & Navigation Publications Maps Comparative Genomics Expression • BLAST • Entrez • Displays • Navigation – extensive crosslinking Phenotypes

  4. NCBI Map Viewer Increasing Discovery Space

  5. NCBI Reference Sequences (RefSeq) • Genome Oriented Resource • A sequence for each macromolecule of Central Dogma • Linked on a residue by residue basis • Objectively non-redundant and comprehensive • Curated Resource • Authoritative source by genome • Derivative of GenBank but corrected, merged, extended • Publicly distributed • Reagents for Genome Annotation and Analysis • Substrate for Functional Genomics

  6. A framework to anchor other information… Disease Structure Function Mature Peptide Development ProPeptide Pathways mRNA Transcript Regulation Chromosome Expression Gene Gene Gene Gene Genetics Organisms Genomes Populations The Basic Model

  7. RefSeq: Scope The NCBI Reference Sequence (RefSeq) project provides non-redundant sequence data including bacterial and viral genomes, mitochondrion, chromosomes, constructed genomic contigs, transcripts, and proteins. RefSeq as a protein database over 280,000 proteins

  8. chromosome NC_000000 protein mRNA gene contig NM_000000 NP_000000 NG_000000 NT_000000 modelmRNA modelprotein XM_000000 XP_000000 RefSeq: Products Goal: One sequence entry for each naturally occurring molecule Multiple products for one gene are instantiated as separate RefSeqs with the same LocusID. Key: curated calculated

  9. Process Flow Database support, automated steps, manual curation New Genes & Descriptions In-house Collaboration LocusLink • Public Release: • LocusLink Web Site BLAST Analysis RefSeq • Provisional & Predicted Records • (Transcripts & Proteins) Status Updates • Reviewed Records • (Genomic Regions,Transcripts, • Proteins) BLAST Curation • Accessible in: • BLAST • BLink • Entrez • FTP • LocusLink Genome Annotation Pipeline

  10. RefSeq: Curation Reviewed Records: Human - 4,685 accessions (3,445 genes) Fly – 1,423 accessions (from FlyBase) Mouse,Rat – 45 accessions Known Genes Gene Families Gene Clusters Identified Problems Assignment Time Review Sequence Database data Literature • Simple cases: • 1-2 days • Large gene families: • months Sequence Editing Correct errors Extend UTRs Splice Variants Annotate Features Final Check Quality Control Public

  11. New Type of RefSeq: Genomic Regions Why? Correct Assembly through Duplications, Paralogous Gene Clusters Optimize Annotation in Gene Clusters Used in Genome Annotation Process

  12. Maintenance New Genes: GenBank UniGene Genome Annotation Collaboration e-mail Updates: GenBank updates Collaboration Ongoing curation Genome Annotation e-mail We welcome feedback, suggestions, collaborations Why Look For RefSeqs? Enhanced Discovery Space: What do we already known? Predicted RefSeqs – where do we need to know more? Genome Annotation Products (Model RefSeqs) Analysis: transcript, protein, annotation, gene index

  13. NCBI Map Viewer Increasing Discovery Space

  14. Gene centered Integrated data RefSeq <-> LocusLink Navigation PubMed OMIM dbSNP RefSeq UniGene GenBank LocusLink Scope: Human Mouse Rat Fly Zebrafish HIV-1

  15. LocusLink: Maintenance Data Collection: Extensive Collaborations with authoritative groups In-house computation In-house curation effort (RefSeq review)

  16. LocusLink: Discovery Find novel uncharacterized genes on a finished chromosome QUERY= 21[chr] NOT has_omim AND has_homol AND type_gene_protein AND predicted AND model AND provisional AND C21orf* OR MGC*

  17. NCBI Map Viewer Increasing Discovery Space

  18. Map Viewer What? Genome Assembly Genome Annotation Integrated map data (genetic, cytogenetic, RH) Scope? Human Drosophila Mouse (model genomes) Why? Facilitate discovery (genes, variation …) Facilitate navigation Facilitate use of genomic sequence information

  19. Genome Build Process STS dbSNP Clones GenomeScan GenBank LocusLink RefSeq Update: Links gi’s Annotation Assembly Resource Updates Input Data: Sequences Curated NTs TPF BLAST hits Public Release Sequences (contig mRNA protein) Map Viewer Analysis & Review Corrections for next build Exclude problem accessions FTP BLAST Input Resources

  20. RefSeq: a reagent for Annotation • Potential Problems: • Gene Families • Partial sequences • Chimeric • Intron read-through • Linker • Vector • Wrong organism genome RefSeq mRNAs GenBank mRNAs • RefSeq Advantages: • Separate Gene Families • Not Partial • Means to correct • problem sequences ESTs TBLASTN RPSBLAST Quality Control: RefSeq review results in excluding problem GenBank sequences from annotation pipeline GenomeScan

  21. What genes (markers, SNPs) are between 2 markers? What BAC clones are available on Xq28? Where are there serine kinases? I’ve cloned gene xyz in my favorite organism. What is related in human? What is the evidence that there is a gene at position n? I have found a phenotype of interest between markers x and y; what is known about this region? What questions can be asked?

  22. Fanconi Syndrome Genetic Mapping Pathology in proximal renal tubular transport

  23. Query Map Viewer Look for genes in the region

  24. Psr2p – sodium stress response in yeast

  25. BLAST Queries: genomic distribution of matches Result from a BLAST query of a zinc finger protein Best match

  26. Review the alignment • A click away: • Alignments (BLAST hit) • Gene Description • Report of all features in the region • Sequence in the region • other mRNAs aligning in the region • Homology Maps • Model Maker - Define your own gene model based on alignments in the region

  27. Anchored by human gene order Anchored by mouse gene order Homology Maps Genes in regions of conserved synteny

  28. Make your own gene model Map Viewer: Model Maker

  29. Entrez GenBank PubMed Expression (GEO) Clones (Clone Registry) NCBI Map Viewer UniGene Markers (UniSTS) HomoloGene Variation (dbSNP) OMIM Homology Maps Increasing Discovery Space

  30. Genome Build Team: Richa Agarwala Hsiu-Chuan Chen Slava Chetvernin Deanna Church Olga Ermolaeva Renata Geer Wratko Hlavina Wonhee Jang Jonathan Kans Ken Katz Paul Kitts David Lipman Adam Lowe Donna Maglott Jim Ostell Kim Pruitt Sergey Resenchuk Victor Sapojnikov Greg Schuler Steve Sherry Andrei Shkeda Tatiana Tatusova Lukas Wagner Sarah Wheelan Acknowledgments RefSeq Curator Staff BLAST Team Entrez Team NCBI Service Desk Staff Collaborators: Human Gene Nomenclature Committee OMIM Staff The Jackson Laboratory Rat Genome Database http://www.ncbi.nlm.nih.gov/