biological databases

Biological Databases

What types of data are available?

What is a database?

What are Genbank and Entrez?

What does a typical entry look like?

How does one use the database?

BIO520 Bioinformatics Jim Lund

NCBI Biological Databases

Central Dogma-o-centric

• Genomic DNA sequence

• mRNA/cDNA sequence

• Protein sequence

• Protein 3D structure

• Literature (Function)

Biological Data• Genomic DNA sequence (complete)• mRNA/cDNA sequence • Gene expression data (NEW)

– Microarrays, SAGE– Expression catalogs

• Protein sequence– Protein interaction/complex data (NEW)

• Protein 3D structure• Literature (Function)

– Organism databases (NEW)– Annotation and classification projects (NEW)

What is a Biological Database?An organized body of persistent data and

associated computer software for updating, querying, and retrieving data records.

• Collection of records and files• Organized for a particular purpose

• The database is separate from the interface and can have several interfaces.– NCBI Protein can be searched by protein name or

using BLAST (Basic Local Alignment Search Tool).

Common database features

• Relational Databases

– Tables

– Relationships between tables

• Version Control

• Consistency enforcement

• Multiauthor/multiuser with security

BIO 520 Student Database

BIO520

Name ID Grade

Amy 123 A

Joe 456 B

Sue 789 C

Table

Record

.

Column

Value

Genbank EntryLOCUS BC005255 495 bp mRNA linear PRI 23-JUN-2006DEFINITION Homo sapiens insulin, mRNA (cDNA clone IMAGE:3950204), complete cds.ACCESSION BC005255VERSION BC005255.1 GI:13528923KEYWORDS MGC.SOURCE Homo sapiens (human) ORGANISM Homo sapiens Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini; Catarrhini; Hominidae; Homo.FEATURES Location/Qualifierssource 1..495 /organism="Homo sapiens"gene 1..495 /gene="INS" /db_xref="GeneID:3630"CDS 60..392 /gene="INS" /translation="MALWMRLLPLLALLALWGPDPAAAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAEDLQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQCCTSICSL YQLENYCN"ORIGIN 1 agccctccag gacaggctgc atcagaagag gccatcaagc agatcactgt ccttctgcca… 421 ccgcctcctg caccgagaga gatggaataa agcccttgaa ccaacaaaaa aaaaaaaaaa 481 aaaaaaaaaa aaaaa//

The CORE: DDBJ, EMBL, and Genbank

Genbank DNA Sequence Database

• Genbank/EMBL/DDBJ mirror & exchange sequence records.

• Primary vs. Secondary Databases– nr (non-redundant database)

• Primary vs. secondary records– Sequence vs. inferred property (coding

region)

Primary vs. Derivative Databases

• Primary Databases– Original submissions by experimentalists– Content controlled by the submitter

• Examples: GenBank, SNP, GEO

• Derivative Databases– Built from primary data– Content controlled by third party (NCBI)

• Examples: Refseq, TPA, RefSNP, UniGene, NCBI Protein, Structure, Conserved Domain

A TraditionalGenBank Record

LOCUS AY182241 1931 bp mRNA linear PLN 04-MAY-2004DEFINITION Malus x domestica (E,E)-alpha-farnesene synthase (AFS1) mRNA, complete cds.ACCESSION AY182241VERSION AY182241.2 GI:32265057KEYWORDS .SOURCE Malus x domestica (cultivated apple) ORGANISM Malus x domestica Eukaryota; Viridiplantae; Streptophyta; Embryophyta; Tracheophyta; Spermatophyta; Magnoliophyta; eudicotyledons; core eudicots; rosids; eurosids I; Rosales; Rosaceae; Maloideae; Malus.REFERENCE 1 (bases 1 to 1931) AUTHORS Pechous,S.W. and Whitaker,B.D. TITLE Cloning and functional expression of an (E,E)-alpha-farnesene synthase cDNA from peel tissue of apple fruit JOURNAL Planta 219, 84-94 (2004)REFERENCE 2 (bases 1 to 1931) AUTHORS Pechous,S.W. and Whitaker,B.D. TITLE Direct Submission JOURNAL Submitted (18-NOV-2002) PSI-Produce Quality and Safety Lab, USDA-ARS, 10300 Baltimore Ave. Bldg. 002, Rm. 205, Beltsville, MD 20705, USAREFERENCE 3 (bases 1 to 1931) AUTHORS Pechous,S.W. and Whitaker,B.D. TITLE Direct Submission JOURNAL Submitted (25-JUN-2003) PSI-Produce Quality and Safety Lab, USDA-ARS, 10300 Baltimore Ave. Bldg. 002, Rm. 205, Beltsville, MD 20705, USA REMARK Sequence update by submitterCOMMENT On Jun 26, 2003 this sequence version replaced gi:27804758.FEATURES Location/Qualifiers source 1..1931 /organism="Malus x domestica" /mol_type="mRNA" /cultivar="'Law Rome'" /db_xref="taxon:3750" /tissue_type="peel" gene 1..1931 /gene="AFS1" CDS 54..1784 /gene="AFS1" /note="terpene synthase" /codon_start=1 /product="(E,E)-alpha-farnesene synthase" /protein_id="AAO22848.2" /db_xref="GI:32265058" /translation="MEFRVHLQADNEQKIFQNQMKPEPEASYLINQRRSANYKPNIWK NDFLDQSLISKYDGDEYRKLSEKLIEEVKIYISAETMDLVAKLELIDSVRKLGLANLF EKEIKEALDSIAAIESDNLGTRDDLYGTALHFKILRQHGYKVSQDIFGRFMDEKGTLE DFLHKNEDLLYNISLIVRLNNDLGTSAAEQERGDSPSSIVCYMREVNASEETARKNIK GMIDNAWKKVNGKCFTTNQVPFLSSFMNNATNMARVAHSLYKDGDGFGDQEKGPRTHI LSLLFQPLVN"ORIGIN 1 ttcttgtatc ccaaacatct cgagcttctt gtacaccaaa ttaggtattc actatggaat 61 tcagagttca cttgcaagct gataatgagc agaaaatttt tcaaaaccag atgaaacccg 121 aacctgaagc ctcttacttg attaatcaaa gacggtctgc aaattacaag ccaaatattt 181 ggaagaacga tttcctagat caatctctta tcagcaaata cgatggagat gagtatcgga 241 agctgtctga gaagttaata gaagaagtta agatttatat atctgctgaa acaatggatt//

Header

Feature Table

Sequence

The Flatfile Format(formatted text)

Genbank Entry

LOCUS PCU30791 1234 bp mRNA PLN 31-MAY-1996

DEFINITION Pneumocystis carinii carinii form 6 guanine nucleotide binding protein alpha subunit (pcg1) mRNA, complete cds.

ACCESSION U30791

NID g1345098

VERSION U30791.1 GI:1345098

Unique IDVersion Control

Content-Taxonomy

SOURCE Pneumocystis carinii f. sp. carinii.

ORGANISM Pneumocystis carinii f. sp. carinii Eukaryota; Fungi; Ascomycota; Archiascomycetes; Pneumocystidaceae; Pneumocystis.

Reference

REFERENCE 1 (bases 1 to 1234)AUTHORS Smulian,A.G., Ryan,M., Staben,C. and Cushion,M. TITLE Signal transduction in Pneumocystis carinii: characterization of the genes (pcg1) encoding the alpha subunit of the G protein (PCG1)

of Pneumocystis carinii carinii and Pneumocystis carinii ratti

JOURNAL Infect. Immun. 64 (3), 691-701 (1996) PUBMED 96186460

•Unique cross reference•Can be >1 reference

Features

FEATURES Location/Qualifiers

source 1..1234 /organism="Pneumocystis carinii f. sp.

carinii“/strain="Form 6“/note="450 kb chromosome" /db_xref="taxon:38081“5'UTR 1..90 gene 91..1155 /gene="pcg1"

CDSCDS 91..1155

/gene="pcg1”

/note="G-protein alpha subunit" /codon_start=1

/product= "guanosine nucleotide binding protein alpha subunit" /protein_id="AAC49295.1"

/db_xref="PID:g1345099" /db_xref="GI:1345099" /translation="MGCCFSATYNQDTLRSKEIE SYLRQEQEHACHEAKILLLGAGES…

.

Related info in another

database

INFERRED

DNA

BASE COUNT 421 a 171 c 195 g 447 t ORIGIN

1 tgaattctaa attttatatt …1201… tattttttta tgctccagat aaaa //

Genbank entries• Combination of required (LOCUS,

SOURCE) and optional fields.– The entry is hierarchical, some fields

contain subfields. • REFERENCE->AUTHORS

• Some fields can appear multiple times (REFERENCE, /gene)

• Some fields are numerical, other are text. Some fields contain free text, others use a controlled vocabulary or an database ID.

Other Genbank output formats

• FASTA– Simple, little annotation information– Easy to use– Common denominator format

• ASN1– Computer friendly, human unfriendly

• XML, INSDSeqXML, TinySeqXML• Graph (graphical map of seq features)…and more

• Genbank (used by VectorNTI)

• FASTA

• GCG– Accelrys GCG (Genetics Computer

Group) package– formerly GCG Wisconsin Package

Many others!

DNA Sequence Files Common formats

FASTA

>gi|1345098|gb|U30791.1|PCU30791 TGAATTCTAAATTTTATATTTCTAATTGCATTTTATATTTTTGATAATACTAGATTTATTCCTGGAAACT TAAATTAGTTATTTTAAGTTATGGGATGTTGTTTTTCTGCTACATATAACCAAGATACACTTCGTTCCAA

One annotation line only!

Submitting sequences to Genbank

•Sequin–Stand-alone sequence submission tool.

•BankIt–Web based sequence submission.

Genbank is an ARCHIVE

•The literature and secondary databases are the knowledge sources.

•There are many additional NCBI annotation databases

NCBI annotation databases!

•Genbank -> RefSeq (Single sequence for each gene)

•Entrez Gene (Gene-based links to annotation sources).•HomoloGene (Homologs)•OMIM•Conserved domains, 3D domains•GEO (Gene expression datasets)•DNA, protein, 3D structures•Interaction data•Links to other databases!

•NCBI Genomes•NCBI Map viewer

Finding and editing DNA files

• Find DNA: Entrez

• Downloading files

• Format Conversion

• Sequence viewing/editing

Entrez• Database

searching/browsing• Example: Pneumocystis G-

proteins– PCR a cDNA to express

in E. coli– Read about it and

related genes– Check similarity to

related G-proteins– View the 3D structure??

•http://www.ncbi.nlm.nih.gov/Entrez/

Entrez Neighbors-Protein

Protein

Protein Literature

BLASTP

citation

DNA

encoding

3D Structurecitation

Mapping the menagerie of biological databases

Nucleic Acid Manipulations

• On the web:– Baylor Human Genome Center (BCM)

http://searchlauncher.bcm.tmc.edu/seq-util/seq-util.html

– European Bioinformatics Institute (EBI) http://www.ebi.ac.uk/Tools/misc.html

DNA/Protein sequence format conversion

• Readseq– Download program:– http://iubio.bio.indiana.edu/soft/molbio/

readseq

– Use online:– http://www.ebi.ac.uk/cgi-bin/readseq.cgi– http://searchlauncher.bcm.tmc.edu/seq-

util/readseq.html

Beware Information Loss!

Reverse Complementing

5’-GAATCA-3’

5’-TGATTC-3’NOT

5’-ACTAAAG-3’

Sequence Statistics

• Nucleotide frequencies (di, tri…)

• UV Absorbance

• MW

• Tm

Restriction Map• Linear vs Circular• Enzyme sets

– Which enzymes, where they cut.

• Gel simulation– Gel-to-map MUCH harder!!

• Useful for:– Cloning– Southern blots– Specialized mol bio techniques

Translation/ORFs

• Translation table– Standard vs non-standard

• Frame (1,2,3,4,5,6)

• Segmental translation (exon-intron)

• Primary translation vs mature polypeptide

Sequence Annotation and Editing

• Text editor– Notepad– Word processor– vi

Nonproportional fonts

(courier, monospaced…)

MWGTCC

IIIIII

MWGTCC

IIIIII

•Artemis•Sequin

•NCBI’s Genbank entry creation/viewing tool

http://frodo.wi.mit.edu/cgi-bin/primer3/primer3_www.cgi

Primer design program: Primer3

Primary vs. Derivative Databases

• Primary Databases– Original submissions by experimentalists– Content controlled by the submitter

• Examples: GenBank, SNP, GEO

• Derivative Databases– Built from primary data– Content controlled by third party (NCBI)

• Examples: Refseq, TPA, RefSNP, UniGene, NCBI Protein, Structure, Conserved Domain

Other NCBI Databases

•Structure: imported structures (PDB)Cn3D viewer, NCBI

curation

•CDD: conserved domain databaseProtein families (COGs

and KOGs)

Single domains (PFAM, SMART, CD)

•dbSNP: nucleotide polymorphism

•Gene: gene recordsUnifies LocusLink and

Microbial Genomes

Homologene Cluster

Entrez Protein: Derivative DatabaseData SourceGenPept

Sequences6,937,176

RefSeq 3,359,561

Third Party Annotation 5,136

Swiss Prot 255,159

PIR 29,996

PRF 12,079

PDB 91,116

PAT Division 669,035

Total 10,690,223

BLAST nr total(no patents or env)

4,545,310

Redundant Proteins

>gi|741682|prf||2007430A DNA mismatch repair protei...MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIV...EDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTAD...

>gi|730028|sp|P40692|MLH1_HUMAN DNA mismatch repair...MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIV...EDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTAD...

>gi|463989|gb|AAC50285.1| DNA mismatch repair prote... MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIV...EDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTAD...

>gi|4557757|ref|NP_000240.1| MutL protein homolog 1...MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIV...EDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTAD...

>gi|13905126|gb|AAH06850.1| MutL protein homolog 1 ... MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIV...EDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTAD...

>gi|1079787|gb|AAA82079.1| DNA mismatch repair prot... MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIV...EDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTAD...

GenPept

NCBI RefSeq

Swiss-Prot

PRF

RefSeq: NCBI’s Derivative Sequence Database

• Curated transcripts and proteins– reviewed– human, mouse, rat, fruit fly, zebrafish, arabidopsis microbial genomes (proteins), and more

• Model transcripts and proteins• Assembled Genomic Regions (contigs)

– human– mouse– rat

• Chromosome records– Human genome– microbial– organelle

ftp://ftp.ncbi.nih.gov/refseq/release/

srcdb_refseq[Properties]

– chicken– honeybee– sea urchin

– zebrafish– cow– dog

– black poplar

RefSeq Accession Numbers

mRNAs and Proteins

NM_123456 Curated mRNANP_123456 Curated ProteinNR_123456 Curated non-coding RNAXM_123456 Predicted mRNAXP_123456 Predicted Protein XR_123456 Predicted non-coding RNAGene RecordsNG_123456 Reference Genomic SequenceChromosomeNC_123455 also Microbial replicons, organelles genomes, human chromosomesAssembliesNT_123456 Contig NW_123456 WGS Supercontig

http://www.ncbi.nlm.nih.gov/RefSeq/key.html