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Sequence Databases – 20 June 2008

Sequence Databases – 20 June 2008. Learning objectives- Be able to describe how information is stored in GenBank. Be able to read a GenBank flat file. Be able to search GenBank for information. Be able to explain the content difference between a header, features and sequence.

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Sequence Databases – 20 June 2008

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  1. Sequence Databases – 20 June 2008 • Learning objectives- • Be able to describe how information is stored in GenBank. • Be able to read a GenBank flat file. • Be able to search GenBank for information. • Be able to explain the content difference between a header, features and sequence. • Be able to say what distinguishes a primary database from a secondary database. • Be able to use and talk about the RefSeq and dbEST databases as they fit into the objectives above. • Be able to access and navigate the ENTREZ platform for biological data analysis.

  2. BIOSEQs – entry common to all sequence databases • BIOSEQ = Biological sequence • Central element in the NCBI database model. • Found in both the nucleotide and protein databases • Comprises the sequence of a single continuous molecule of nucleic acid or protein. Entry must have • At least one sequence identifier (Seq-id) • Information on the physical type of molecule (DNA, RNA, or protein) • Descriptors, which describe the entire Bioseq • Annotations, which provide information regarding specific locations within the Bioseq

  3. What is GenBank? • The NIH genetic sequence database, an annotated collection of all publicly available NUCLEIC ACID sequences • Each record represents a single contiguous stretch of DNA or RNA • DNA stretches may have more than one coding region (i.e., more than one gene). • RNA sequences are presented with T, not U • Records are generated from direct submissions to the DNA sequence databases from the investigators (authors). • GenBank is part of the International Nucleotide Sequence Database Collaboration.

  4. The number of basepairs is now at over 85 billion. The number of sequences is approaching 83 million.

  5. General Comments on GBFF • Three sections: • 1) Header-information about the whole record • 2) Features-description of annotations-each represented by a key. • 3) Nucleotide sequence-each ends with // on last line of record. • Nucleic acid (DNA or RNA (cDNA)) sequence translated to amino acid sequence is a “feature” • Genbank Flat File (MyoD1 as an example)

  6. Feature Keys • Purpose: • 1) Indicates biological nature of sequence • 2) Supplies information about changes to sequences Feature KeyDescription conflict Separate determinations of the same seq. differ rep_origin Origin of replication protein_bind Protein binding site on DNA CDS Protein coding sequence

  7. Feature Keys-Terminology Feature Key Location/Qualifiers CDS 23..400 /product=“alcohol dehydro.” /gene=“adhI” The feature CDS is a coding sequence beginning at base 23 and ending at base 400, has a product called “alcohol dehydrogenase” and corresponds to the gene called “adhI”.

  8. Feature Keys-Terminology (Cont.) Feat. Key Location/Qualifiers CDS join (544..589,688..1032) /product=“T-cell recep. B-ch.” /partial The feature CDS is a partial coding sequence formed by joining the indicated elements to form one contiguous sequence encoding a product called T-cell receptor beta-chain. (For MyoD1 – Accession number X61655)

  9. Record from GenBank GenBank division (plant, fungal and algal) Locus name Modification date LOCUS SCU49845 5028 bp DNA PLN 21-JUN-1999 DEFINITION Saccharomyces cerevisiae TCP1-beta gene, partial cds, and Axl2p (AXL2) and Rev7p (REV7) genes, complete cds. ACCESSION U49845 VERSION U49845.1 GI:1293613 KEYWORDS . SOURCE baker's yeast. ORGANISM Saccharomyces cerevisiae Eukaryota; Fungi; Ascomycota; Hemiascomycetes; Saccharomycetales; Saccharomycetaceae; Saccharomyces. Unique identifier (never changes) Coding region GeneInfo identifier (changes whenever there is a change) Nucleotide sequence identifier (changes when there is a change in sequence (accession.version)) Word or phrase describing the sequence (not based on controlled vocabulary). Not used in newer records. Common name for organism Formal scientific name for the source organism and its lineage based on NCBI Taxonomy Database

  10. Record from GenBank (cont.1) REFERENCE 1 (bases 1 to 5028) AUTHORS Torpey,L.E., Gibbs,P.E., Nelson,J. and Lawrence,C.W. TITLE Cloning and sequence of REV7, a gene whose function is required for DNA damage-induced mutagenesis in Saccharomyces cerevisiae JOURNAL Yeast 10 (11), 1503-1509 (1994) MEDLINE 95176709 REFERENCE 2 (bases 1 to 5028) AUTHORS Roemer,T., Madden,K., Chang,J. and Snyder,M. TITLE Selection of axial growth sites in yeast requires Axl2p, a novel plasma membrane glycoprotein JOURNAL Genes Dev. 10 (7), 777-793 (1996) MEDLINE 96194260 Medline UID REFERENCE 3 (bases 1 to 5028) AUTHORS Roemer,T. TITLE Direct Submission JOURNAL Submitted (22-FEB-1996) Terry Roemer, Biology, Yale University, New Haven, CT, USA Submitter of sequence (always the last reference)

  11. Record from GenBank (cont.2) There are three parts to the feature key: a keyword (indicates functional group), a location (instruction for finding the feature), and a qualifier (auxiliary information about a feature) FEATURES Location/Qualifiers source 1..5028 /organism="Saccharomyces cerevisiae" /db_xref="taxon:4932" /chromosome="IX" /map="9" CDS <1..206 /codon_start=3 /product="TCP1-beta" /protein_id="AAA98665.1" /db_xref="GI:1293614" /translation="SSIYNGISTSGLDLNNGTIADMRQLGIVESYKLKRAVVSSASEA AEVLLRVDNIIRARPRTANRQHM" Location Keys Qualifiers The 5’ end of the coding sequence begins upstream of the first nucleotide of the sequence. The 3’ end is complete. Start of open reading frame Descriptive free text must be in quotations Database cross-refs Protein sequence ID # Values Note: only a partial sequence

  12. Record from GenBank (cont.3) Another location gene687..3158 /gene="AXL2" CDS 687..3158 /gene="AXL2" /note="plasma membrane glycoprotein" /codon_start=1 /function="required for axial budding pattern of S. cerevisiae" /product="Axl2p" /protein_id="AAA98666.1" /db_xref="GI:1293615" /translation="MTQLQISLLLTATISLLHLVVATPYEAYPIGKQYPPVARVN. . . “ gene complement(3300..4037) /gene="REV7" CDS complement(3300..4037) /gene="REV7" /codon_start=1 /product="Rev7p" /protein_id="AAA98667.1" /db_xref="GI:1293616" /translation="MNRWVEKWLRVYLKCYINLILFYRNVYPPQSFDYTTYQSFNLPQ . . . “ Cutoff Another location Cutoff

  13. Record from GenBank (cont.4) BASE COUNT 1510 a 1074 c 835 g 1609 t ORIGIN 1 gatcctccat atacaacggt atctccacct caggtttaga tctcaacaac ggaaccattg 61 ccgacatgag acagttaggt atcgtcgaga gttacaagct aaaacgagca gtagtcagct . . .//

  14. Primary databases vs. Secondary databases • Primary database • comprises information submitted directly by the experimenter. • is called an archival database. • Secondary database • comprises information derived from primary database. • is a curated database.

  15. NCBI site map • To notice on the map • General organization • Where the following fit: • RefSeq (nucleotide, protein) • dbEST • Others of interest to you • NCBI site map: http://www.ncbi.nlm.nih.gov/Sitemap/index.html

  16. GenBank/EMBL/DDBJ PDB-Three-dimensional structure coordinates of biological molecules PROSITE-database of protein domain/function relationships. http://www.expasy.org/prosite/ Types of primary databases carrying biological infomation

  17. Protein databases derived from GenBank containing data for a single gene • Non-redundant (nr) • Swissprot • PIR (Int’l. protein sequence) • LocusLink • RefSeq Secondary Databases protein DNA RNA cDNA DNA databases derived from GenBank containing data for a single gene • Non-redundant (nr) • dbGSS (genome survey sequences) • dbHTGS (high throughput) • dbSTS (sequence tagged site) • LocusLink • RefSeq RNA (cDNA) databases derived from GenBank containing data for a single gene • dbEST (expressed sequence tag) • UniGene • LocusLink • RefSeq

  18. RefSeq-Comprehensive, integrated, non-redundant, well-annotated set of sequences, including genomic DNA, transcripts, and proteins. http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=15608248 Types of secondary databases carrying biological infomation

  19. Some nucleotide secondary databases dbEST- Sequence data and other information on "single-pass" cDNA sequences, or "Expressed Sequence Tags", from a number of organisms Genome databases-(there are over 20 genome databases that can be searched) EPD:eukaryotic promoter database http://www.epd.isb-sib.ch/ NR-non-redundant GenBank+EMBL+DDBJ+PDB. Entries with 100% sequence identity are merged as one. Types of secondary databases carrying biological infomation

  20. Some protein secondary databases ProDom http://protein.toulouse.inra.fr/prodom/current/html/home.php PRINTS http://bioinf.man.ac.uk/dbbrowser/PRINTS/ BLOCKS http://bioinformatics.weizmann.ac.il/blocks/ Types of secondary databases carrying biological infomation

  21. References for understanding the NCBI sequence database model • Here is the website for NCBI developer tools. • http://www.ncbi.nlm.nih.gov/IEB/ToolBox/SDKDOCS/INDEX.HTML

  22. RNA, but NOT mRNA RNA, but NOT mRNA Mature mRNA DNA  RNA PROTEINRNA processing

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