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Using Entrez The Life Sciences Search Engine. Searching NCBI Databases Efficiently Knowing how to retrieve the exact information you need in an efficient.

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Presentation on theme: "Using Entrez The Life Sciences Search Engine. Searching NCBI Databases Efficiently Knowing how to retrieve the exact information you need in an efficient."— Presentation transcript:

1 Using Entrez The Life Sciences Search Engine

2 Searching NCBI Databases Efficiently Knowing how to retrieve the exact information you need in an efficient way is the fundamental and most important skill in Bioinformatics. Every NCBI database is designed and created for some specific purposes. A common mistake Bioinformatics novices make is searching for information in an inappropriate database. Entrez links among and within databases, making it easier to search for information.

3 What is Entrez? Entrez is an NCBI retrieval system designed for searching several linked databases. Entrez is a search tool for integrated access to the biological literature and sequence data. Entrez is extremely powerful, enabling the user to quickly move between the different specialized databases.

4 Entrez Entrez is divided into sites for nucleotide, protein, structure, genomes, OMIM, and more. You can use limits (such as RefSeq) to focus your Entrez search. When you conduct a search via Entrez, your query generates this screen, telling you the number of hits to your query.

5 The Entrez System

6 The Big Picture LocusLink Nucleotide Protein OMIM PubMed SNP MGC UCSC GDB e! HGMD UniGene Homologene MapViewer Structure 3D Domains CDD Books PopSet Genome Taxonomy ProbeSet UniSTS Entrez

7 Entrez and LocusLink Entrez doesn’t link to all the databases that contain sequences, however! LocusLink has its own groups of links to specialty databases, since it doesn’t cover all the genomes yet.

8 Genomes Taxonomy Entrez: Database Integration PubMed abstracts Nucleotide sequences Protein sequences 3-D Structure Word weight VAST BLAST Phylogeny

9 Entrez Journals UniGene PubMedNucleotide Protein SNP Genome BooksProbeSet OMIM CDD Taxonomy 3D Domains UniSTS PopSet Structure The (ever) Expanding Entrez System

10 Entrez Databases PubMedBiomedical literature BooksOnline textbooks NucleotideGenBank, EMBL, DDBJ, RefSeq, PDB Protein[GenBank, EMBL, DDBJ], RefSeq, SWISS-PROT, PIR, PRF, PDB GenomeComplete genomes TaxonomyOrganisms in NCBI sequence databases StructureMMDB: experimental 3D structures DomainsCDD: conserved protein domains 3D DomainsCompact 3D protein domains in MMDB OMIMOnline Mendelian Inheritance in Man SNPSingle nucleotide polymorphisms UniSTSSequence Tagged Site markers ProbeSetGene expression and microarray datasets PopSetPopulation study datasets UniGeneGene-based expressed sequence clusters

11 Nucleotide Database The Nucleotide database contains sequence data from GenBank, EMBL, and DDBJ, the members of the tripartite, international collaboration of sequence databases. EMBL is the European Molecular Biology Laboratory at Hinxton Hall, UK; DDBJ is the DNA Database of Japan in Mishima, Japan. Sequence data are also incorporated from the Genome Sequence Data Base (GSDB), Santa Fe, NM. Patent sequences are incorporated through arrangements with the U.S. Patent and Trademark Office (USPTO) and via the collaborating international databases from other international patent offices.

12 Entrez Nucleotides Primary GenBank / EMBL / DDBJ 35,116,960 Derivative RefSeq 259,219 Third Party Annotation 3,182 PDB 4,703 Total 35,384,248

13 Database Searching with Entrez uUsing limits and field restriction to find plant g6pdh uLinking and neighboring with g6pdh

14 Entrez Nucleotides glucose 6 phosphate dehydrogenase The G6PD enzyme catalyzes the oxidation of glucose-6- phosphate to 6-phosphogluconate, while reducing nicotinamide adenine dinucleotide phosphate (NADP+ to NADPH). In terms of electron transfer, glucose-6-phosphate loses two electrons to become 6-phosphogluconate and NADP+ gains two electrons to become NADPH. This is the first step in the pentose phosphate pathway. This pathway, or shunt, as it is sometimes called, produces the 5- carbon sugar, ribose, which is an essential component of both DNA and RNA.

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16 Limits Are Helpful Limits allow restriction of a search to a defined subset of the database. Limits can be set to restrict a search to a particular database field (e.g., the Author field). Limits can be set to search everything but a particular type of data (e.g., “exclude patent records”). Alternatively, limits can be set to search only a particular type of data (e.g., Genomic RNA/DNA) or to search only data from a particular source database (e.g., EMBL). Date limits and sequence length limits are also possible. The contents of each Entrez database differ, and therefore the Limits available for each database differ.

17 glucose 6 phosphate dehydrogenase Entrez Nucleotides: Limits & Preview/Index Try using the Limits and Preview function to hone your search To find the Plant G6PD genes.

18 glucose 6 phosphate dehydrogenase Entrez Nucleotides: Limits Accession All Fields Author Name EC/RN Number Feature key Filter Gene Name Issue Journal Name Keyword Modification Date Organism Page Number Primary Accession Properties Protein Name Publication Date SeqID String Sequence Length Substance Name Text Word Title Word Uid Volume Field Restriction Exclude bulk sequences

19 glucose 6 phosphate dehydrogenase Entrez Nucleotides: Limits Title == Definition Exclude Bulk Sequences mRNA molecule type Nuclear gene

20 Document Summaries: Limits

21 green plants Adding Terms: Preview/Index Accession All Fields Author Name EC/RN Number Feature key Filter Gene Name Issue Journal Name Keyword Modification Date Organism Page Number Primary Accession Properties Protein Name Publication Date SeqID String Sequence Length Substance Name Text Word Title Word Uid Volume green plants

22 Plant cytosolic g6pdh mRNAs

23 Database Neighbors and Interlinking What makes Entrez more powerful than many services is that most of its records are linked to other records, both within a given database (such as Nucleotide) and between databases. Links within a database are called “neighbors” (e.g., Nucleotide neighbors).

24 Links Between Databases Protein and Nucleotide neighbors are determined by performing similarity searches using the BLAST algorithm to compare the entry amino acid or DNA sequence to all other amino acid or DNA sequences in the database. We will discuss more about BLAST later. Nucleotide sequence records in the Nucleotide database are linked to the PubMed citation of the article in which the sequences were published. Protein sequence records are linked to the nucleotide sequence from which the protein was translated.

25 Plant cytosolic g6pdh mRNAs Summary Brief GenBank ASN.1 FASTA GI list LinkOut PubMed Links Protein Links Nucleotide Neighbors PopSet Links Structure Links Genome Links Taxonomy Links OMIM Links Formats Links and neighbors (related records)

26 LinkOut LinkOut is a feature of Entrez that is designed to provide users with links from PubMed and other Entrez databases to a wide variety of relevant web- accessible online resources: –Full-text publications –Other biological databases –Consumer health information –Research tools The goal is to facilitate access to relevant online resources beyond the Entrez system to extend, clarify, or supplement information found in the Entrez databases.

27 Protein Database The protein database includes proteins from translate regions of DNA in GenBank as well as sequence from PIR The entry includes: –The name of the protein – How the protein sequence was derived –An accession and a PID number –The number of amino acids

28 Protein Entry The Entry also includes: Structural information for the protein (if known) –Helices and  - Sheets –Domains –Etc The sequence of amino acids comprising the protein

29 Setting Protein Database search limits Choose Protein from the drop-down menu –Can do a Boolean search –Or can set LIMITS Fields (eg Author, Journal, etc.) Gene Location (genomic, mitochondrial etc) Segmented Sequence Only from (Database to check) Modification date

30 Linking Between Databases Sometimes you will pull up a record and you have no idea what organism the gene you are looking at is from. For Example, the following record- what is Medicago sativa ?

31 Entrez GenBank / GenPept

32 Taxonomy to the Rescue Entrez lets you click a live link from the record and determine what organism Medicago sativa is. It is alfalfa. You can also tell what it is related to taxonomically, because sometimes the common name isn’t very useful either!

33 Taxonomy Link

34 Advanced Neighbors: BLink

35 What is BLink BLink - BLAST Link Someone has done a BLAST search already, and you can just retrieve it! BLink displays the graphical output of pre- computed blastp results against the protein non-redundant (nr) database.

36 This graphical output includes: Alignment of up to 200 BLAST hits on the query sequence Best Hits to each organism List of known protein domains in the query sequence Filter hits by selecting the BLAST cutoff score Distribution of hits by taxonomic grouping Display of similar sequences with known 3D structure Filter hits by database and/or by taxonomic grouping Display a taxonomic tree of all organisms with similar sequences

37 PopSet Links The PopSet database contains aligned sequences submitted as a set resulting from a population, phylogenetic, or mutation study. These alignments describe such events as evolution and population variation. The PopSet database contains both nucleotide and protein sequence data.

38 Protein Neighbors->PopSet Links

39 Protein Neighbors->Genome Links

40 PopSet search results The results or a PopSet search The PopSet database includes alignments of genes from multiple organisms OR different gene families OR mutational analyses

41 PopSet Entry The PopSet entry includes: –The title of the paper/study –The length of the sequence(s) aligned –The number of aligned sequences

42 PopSet Entry without alignment The PopSet Entry without an alignment –Title of the study –The number of sequences included –Links to the sequences

43 Entrez Structures

44 Protein Structures can also be in databases is a useful review Tutorial.

45 Entrez links to structure databases The Structure database or Molecular Modeling Database (MMDB) contains experimental data from crystallographic and NMR structure determinations. The data for MMDB are obtained from the Protein Data Bank (PDB). The NCBI has cross-linked structural data to bibliographic information, to the sequence databases, and to the NCBI taxonomy. Use Cn3D, the NCBI 3D structure viewer, for easy interactive visualization of molecular structures from Entrez.

46 Structure Search results The structure of proteins are also in a database Search as before Your search results are similar

47 Structure Entry The structure Entry has links to the other databases And it will allow you download a file to open with a structure viewer program

48 Proteins with similar structures and functions have been identified in the databases

49 BLink: Advanced Protein Neighbors

50 BLink: Related Structures

51 Viewing Structure in Cn3D You can download Cn3D (a structural viewer program) from NCBI This will allow you to view the structures from the structure database

52 Cn3D Text Window The Text window of Cn3D will align two or more proteins so you can compare the structure of multiple proteins

53 BLink: Human Homologue

54 Human RefSeqs: Genome Reagents

55 MM MMDB: Molecular Modeling Data Base Derived from experimentally determined PDB records Value added to PDB records including: –Addition of explicit chemical graph information –Validation –Inclusion of Taxonomy, Citation, and other information –Conversion to ASN.1 data description language Structure neighbors determined by Vector Alignment Search Tool (VAST)

56 Structure Summary Cn3D viewer Conserved Domains 3D Domain Neighbors Structure Neighbors

57 Cn3D 4.1

58 Cn3D 4.1: Structural Alignment Casein kinase S. pombe Src Kinase H. sapiens Conserved ATP binding site

59 Cn3D: Simple Homology Modeling human swordtail

60 Using Cn3D to model domains

61 Other services and databases from the NCBI LocusLink to all possible information from NCBI and beyond for a few well characterized model organisms. LocusLink is a great starting point: it collects key information on each gene/protein from major databases. It now covers 8 organisms. RefSeq provides a curated, optimal accession number for each DNA (NM_006744) or protein (NP_007635)

62 Locus Links Results of a Locus links search, includes: –Locus ID –Species –Locus symbol –Locus name –Locus location –Links P rotein Database O MIM R eference Sequence R elated GenBank Sequences H omologene Data U niGene V ariation Data

63 LocusLink: Selected Higher Genomes OMIM RefSeq GenBank dbSNP UniGene Full report PubMed HomoloGene Map Viewer Protein

64 Protein Database The Protein database contains sequence data from the translated coding regions from DNA sequences in GenBank, EMBL, and DDBJ as well as protein sequences submitted to: –Protein Information Resource (PIR) –SWISS-PROT –Protein Research Foundation (PRF) –Protein Data Bank (PDB) (sequences from solved structures)

65 NCBI Protein Databases GenPept GenBank, EMBL, DDBJ CDS translations RefSeq mRNA based (NP_) and genome based (XP_) Swiss-Prot curated high quality protein reviews PIR protein information resource Georgetown University PRF protein resource foundation PDB Protein Databank sequences from structures

66 Entrez Protein GenPept (GB,EMBL, DDBJ) 3,442,298 RefSeq 856,191 Third Party Annotation 3,834 Swiss Prot 144,508 PIR 282,821 PRF 12,079 Total 3,442,298 BLAST nr 1,642,191

67 Protein Link BLAST Link Conserved Domains

68 Related Proteins: Redundancy Redundant Sequences

69 Sequence from MutL structure Related Proteins: Links

70 BLink: non-redundant relatives Arabidopsis homolog Conserved Domain

71 MLH1 Domain Structure: CDD ATPase Domain Mismatch Repair Domain

72 MLH1: ATPase Domain

73 1BGQ: ATPase Domain in Cn3D Yeast HSP90 ATP Binding site helix

74 Variations Human MLH1

75 BLink Finding structural models

76 Mapping Variation Onto Structure Bacterial DNA mismatch repair proteins Loads sequence alignment and structure in Cn3D

77 Mapping Variation Onto Structure Conserved Asn Asn Ile Ile – Val

78 NCBI Genome Databases The Genome database provides views for a variety of genomes, complete chromosomes, sequence maps with contigs, and integrated genetic and physical maps.

79 Microbial Genomes ZWF

80 Genome search results Genome Search Results The Genome database includes full (and some partial) genomes from viruses to complex organisms

81 Genome Entry Genome entries include –Maps of the genome –Links to the sequence –The organism for the genome

82 Genes Database: All Genomes Coming soon!

83 Genes Database: All Genomes

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85 But wait! There’s more! There is even more at NCBI that I have covered here. This site map is also a guide to NCBI resources. Each link leads to a brief description of the resource on this page, then to the resource itself.

86 There are many bioinformatics servers outside NCBI. Try ExPASy’s sequence retrieval system at (ExPASy = Expert Protein Analysis System) Or try ENSEMBL at for a premier human genome web browser.


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