1. Retrieving Information: Using Entrez
B.F. Francis Ouellette
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2. Before we get started Curated vs non-curated Databases
Intellectual value added by curation process.
Updated of record, a case study:
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3. Updates -- the 4 “W” Who updates? Submitters, Journals, “3rd party”
What to update? Gene names, citations, new product, sequencing errors
Where?
Why update?
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4. example
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7. To: ddbjupdt@ddbj.nig.ac.jp Subject: D25291 mito Dear colleagues,
From francis Wed Mar 3 22:32:
To:
Subject: D25291 mito
Dear colleagues,
it appears that DDBJ record D25291 is contaminated with mitochondrial
sequences from nucleotide 673 to 1803, as it is identical to mouse
mitochondrial sequence (EMBL V00711) for more than 1100 nucleotides.
I would recommend deleting that segment of the record, or removing the
record altogether, as it leads to unfortunate misinterpretation of the
data when using GenBank or DDBJ. The protein sequence (which is
erroneous, as it is all of mitochondrial origin) should definitely be
removed as well. ….
LOCUS MUSNGH bp mRNA ROD AUG-1997
DEFINITION Mouse neuroblastoma and rat glioma hybridoma cell line NG108-15
cell TA20 mRNA, complete cds.
ACCESSION D25291
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8. Sequence Updated length Date DEF Version GI
LOCUS MUSNGH bp mRNA ROD AUG-1997
DEFINITION Mouse neuroblastoma and rat glioma hybridoma cell line NG108-15
cell TA20 mRNA, complete cds.
ACCESSION D25291
NID g
VERSION D GI:
length
Date
DEF
Version GI
LOCUS MUSNGH bp mRNA ROD MAR-1999
DEFINITION Mouse neuroblastoma and rat glioma hybridoma cell line NG108-15
mRNA.
ACCESSION D25291
VERSION D GI:
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9. Another guiding principals when using GenBank
Data in GenBank is only as good as what you put in: applying and ensuring this (in an active, day to day fashion) will only make everybody’s work that much easier.
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10. Nature 2001 Jan 25; 409:452
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11. Check sequence revision history
Nucleotide Go
Check sequence revision history
Check sequence revision history
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12. Check the history for D25291
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14. Back to Today’s lecture
Using Entrez …
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15. Retrieving information: how it works:
Servers have the records you want
You need to understand the data they have, and how it is organized
There are often many ways to get to an answer.
Route to get there is not always obvious, but you need to think of alternatives and traps.
Use some query language – each system has its own.
Retrieve data in a specified format.
Save it in a way that will be useful to you.
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16. What you may be looking for:
Did a BLAST search – and you need more info about some of the proteins they found similarities to.
Heard on CBC about a disease gene that was recently discovered, and you want to know more about it.
Want to build a dataset for local blast searches.
A colleague wants you to do an alignment of all sequences from a given protein family.
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17. What you are looking for:
PubMed paper from author X
Sequence from gene X in organism Y
All information about organelle W in model organism Y
All information about disease X in human
Orthologs of that disease genes in other model organisms
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18. Central Dogma in Biology
DNA
RNA
The central dogma in Biology:
DNA makes DNA and also makes RNA which makes proteins
protein
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19. Central Dogma: NCBI version
DNA
RNA
Write a paper
about it
protein
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20. Entrez: Pathway to Discovery
Term frequency statistics
1993
MEDLINE abstracts
Literature citations in sequence databases
Literature citations in sequence databases
Nucleotide sequences
Protein sequences
Nucleotide sequence similarity
Amino acid sequence similarity
Coding region features
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21. Type in your last name and find a paper form one of your teammates
Related Articles
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22. Hard link DNA to protein L12345
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23. 2003 From Fig 1 of Entrez search and retrieval system Jim Ostell
Chapter 14, the NCBI Handbook.
2003
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27. Ctrl-F
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29. Getting started in Entrez
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30. A query: All the yeast papers by the instructor of this lecture
1st challenge: how do you spell his name again? What are his initials?
Do I need to know the Greek name for yeast? Which yeast?
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31. “ouellette bf” [au] AND yeast
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35. MeSH: Medical Subject Heading
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36. A query Word <free text> : too many hits
More words (the Boolean ‘AND’ is the default)
Limit query to specified field
Limit query in time
Do Boolean on queries
#1 AND #2
#3 NOT #5
#7 OR #8
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37. hieter p [au]
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38. Limit in Time:
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40. Full Text in PubMed Central
No abstract
With abstract
Full Text on-line
Full Text in PubMed Central
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41. boguski m [au]
boguski ms [au]
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42. #24 NOT #
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44. Other types of links in Entrez
Next slides to explore other kind of things linked into Entrez records.
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45. “hieter p” [au] cdc16p
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54. “Books”
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55. (2)
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61. Link to Genome View of Chromosome I
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64. RefSeq
RefSeq represents the NCBI curated “reference sequences” for all ‘worked’ genome.
Historically, these used to be referred to as “GenBank-Gold”.
RefSeq are either genomic, mRNA or protein sequences.
Not all sequences are in RefSeq
All RefSeq sequences are assembled/taken from things in GenBank.
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65. Some of the features of the RefSeq:
 non-redundancy  
explicitly linked nucleotide and protein sequences  
updates to reflect current knowledge of sequence data and biology  
data validation and format consistency  
distinct accession series  
ongoing curation by NCBI staff and collaborators, with review status indicated on each record
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66. Accession number space
GenBank:
1+5 (L12345, U00001)
2+6 (AF000001, AC000003)
4+2+6 (WGS)
All have accession.version
Protein:
1+5 (SwissProt/UniProt)
3+5 (GenPept)
RefSeq:
N*_12345
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67. RefSeq Accession Number Space
NC_123456
Genomic
Complete genomic molecules including genomes, chromosomes, organelles, plasmids.
NG_123456
Incomplete genomic region; supplied to support the NCBI Genome Annotation pipeline.
NM_123456
mRNA
NR_123456
RNA
Non-coding transcripts including structural RNAs, transcribed pseudogenes, and others
NP_123456
Protein
NP_
Planned expansion of accession series
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68. Automated Assemblies NT_123456 Genomic
Intermediate genomic assemblies of BAC sequence data
NW_123456
Intermediate genomic assemblies of Whole Genome Shotgun sequence data
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69. Model RefSeq records XM_123456 mRNA
model mRNA provided by the Genome Annotation process; sequence corresponds to the genomic contig.
XR_123456
RNA
model non-coding transcripts provided by the Genome Annotation process; sequence corresponds to the genomic contig.
XP_123456
Protein
model proteins provided by the Genome Annotation process; sequence corresponds to the genomic contig.
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70. WGS special case NZ_ABCD12345678 Genomic
A collection of whole genome shotgun sequence data for a project. Accessions are not tracked between releases. The first four characters following the underscore (e.g. 'ABCD') identifies a genome project.
ZP_
Protein
Proteins annotated on NZ_ accessions (often via computational methods).
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71. Download all the data
Entrez and RefSeq
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75. Locus Link
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76. Things to watch out for:
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78. In-Lab Exercise Questions for this exercise in Binder
Work with your teammates
Feel free to explore the information space
In PubMed look up your favorite author, people you went to school with, or people at an institution you would like to work at some day!
This Lab is NOT marked
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