Sequence Analysis Unit 5

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Sequence AnalysisUnit 5

• BIOL221T Advanced Bioinformatics for
  Biotechnology

Irene Gabashvili, PhD igabashvili_at_yahoo.com
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Reminder

• Lectures Lab every Wednesday, Duncan Hall,
  Room 550, 600 pm to  945 pm
• Office hours Wednesday, 4pm-6pm (Room 554,
  phone [phone]) and by appointment
• Lecture notes http//home.comcast.net/igabashvil
  i/221T.htm
• Or the SJSU page --
• The user name is ewok\biostudents (dont enter
  quotation mark)  
• And the password is 4biolecture (dont enter
  quotation mark).

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Problem Set Review

• 1. SWISSPROT protein sequence database began in
  1987 all correct answers
• 2. OMIM stands for "Online Mendelian Inheritance
  in Man all correct answers
• 3. How many genes in human 6q23 region?
• 6q23 AND humanorgn (AND aliveprop)
  (EntrezGene) ? 320(76) (6q23.1-6q23.3)
• 11 genes exactly in 6q23 (decimal not known)
• MapViewer 1505 genes on chr.6, 72 between
  130,400K 139,100K, 93 between 23.1 and 23.3

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Why different results?
Entrez Gene
6q23 AND humanorgn AND aliveprop
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Entrez Map Viewer

• Good tools, but lots of bugs be aware
• Make sure the right numbers are displayed in the
  Region Shown boxes the software tends to null
  them and display something else. If nothing is
  shown wrong results! If different numbers are
  shown wrong results! Only if the correct
  numbers/loci are still there, trust the results.

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Map View Allowable Values

• cytogenetic bands - if the master map is a
  cytogenetic map, you can enter band numbers in
  formats such as 9p23 to 9p11, or 22q12.1 to
  22q13.2.
• symbols - gene symbols, marker names, etc to
  display a region of the chromosome between
• numerical positions -cM for genetic linkage map,
  cR for radiation hybrid map, base pairs - if the
  master map is a sequence map1000000 or
  1,000,000 or 1M or 1000K or 1.1M or 1.2K

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Entrez Map Viewer

• Types of maps might include
• cytogenetic map (banding pattern) also referred
  to as ideogram
• genetic linkage map (centiMorgans cM)
• radiation hybrid map (centiRays cR)
• sequence map (megabases Mb)

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• http//www.ncbi.nlm.nih.gov/mapview/maps.cgi?taxid
  9606chr7mapsideogrverboseon

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Why different results?

• OMIM lists not only known genes but also regions
  associated with diseases e.g.
• Pulmonary function 6q21-q22
• Stature (height) 6q24-q25
• Even if the answer appears right, it might
  include a few extra genes -- 6q24-q25
• http//www.ncbi.nlm.nih.gov/Omim/getmap.cgi?chromo
  some6q23
• Make sure you use the right maps!

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Entrez Map Viewer

• Where does gene X exist within the genome of
  organism Y? What are some flanking markers?
• Which genes exist on a chromosome, and in what
  order do they appear?
• Show the genes that exist in region R, the
  corresponding sequence data for that region.
• Display the region of a chromosome between points
  A and B.
• I know the cytogenetic location of gene X. What
  is the corresponding physical location?

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genome.ucsc.edu UCSC Genome Browser
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Problem Set Review

• 4. What gene has SNP rs3761847?
• http//www.ncbi.nlm.nih.gov/projects/SNP/
• 5. OMIM Hypertensiondis AND (KCNA1gene OR
  TRPM3gene OR NOS3gene OR CHEK2gene)
• 6. clinicaltrials.gov 6
• 7. crisp.cit.nih.gov 1400
• 8. EST

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Alignment
NT_017582.2
C5 TRAF1 PRO1995
TNF receptor associated factor 1
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Problem Set Review

• 9. 90000100000SLEN ? 32K records
• 10. txid9606 NOT newentrytitle NOT aliveprop
  ? 126K
• 11. qualifier to get less results gene
• 12. Gene, OMIM
• 13. KEGG pathway Histidine metabolism 00340 KEGG
  pathway Pentose phosphate pathway 00030 KEGG
  pathway Purine metabolism 00230 Reactome Event
  Metabolism of carbohydrates 71387
• 14. Yes, Entrez Gene DB provides info on
  protein-protein interactions, if known

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BaxevanisOuellette ch.1

• Intro
• Primary and Secondary DBs
• Nucleotide Sequence DBs
• DB formats
• Nucleotide Sequence Flatfiles
• The feature table
• Third Party Annotation

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Sequence Databases

• 1965 M.Dayhoffs Atlas of Protein Sequences
• 1979 - Los Alamos Sequence Library (pre-GenBank)
• 1980 EMBL, 1st public nucleotide sequence DB
• 1982 GenBank (formerly Los Alamos Library)
• 1984 DDBJ - DNA Data Bank of Japan
• 1988 - NCBI created at NIH/NLM
• 1991 the 1st version of Entrez distributed on
  CD-ROM
• 1992 GenBank transferred to NCBI

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Links

• http//www.ncbi.nlm.nih.gov/Genbank/
• http//www.ebi.ac.uk/embl/
• http//www.ddbj.nig.ac.jp/
• The three databases exchange data daily. For
  formats, codes, etc, see feature table
• http//www.ncbi.nlm.nih.gov/collab/FT/index.html

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Submitting to these DBs

• Sakura, a nucleotide sequence data submission
  system http//sakura.ddbj.nig.ac.jp/
• Sequin, submitting to GenBank www.ncbi.nlm.nih.go
  v/Sequin
• Submitting to EMBL www.ebi.ac.uk/embl/Submission/
  webin.html

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About

• http//www.ncbi.nlm.nih.gov/books/bv.fcgi?ridhand
  book.chapter.ch1 (GenBank, e-book)
• http//nar.oxfordjournals.org/cgi/content/full/36/
  suppl_1/D25 (GenBank, NAR-2008)
• http//nar.oxfordjournals.org/cgi/content/full/36/
  suppl_1/D22 (DDBJ, NAR-2008)
• http//nar.oxfordjournals.org/cgi/content/full/35/
  suppl_1/D16 (EMBL, NAR-2007)

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Pubmed Nucleic Acids Res.journal AND "Database
issue"

• PRIDE,  PRoteomics IDEntifications database
• http//www.ebi.ac.uk/pride/
• STITCH, Chemical-Protein Interactions
• http//stitch.embl.de/
• The universal protein resource (UniProt).
  http//www.uniprot.org
• http//nar.oxfordjournals.org/cgi/content/full/36/
  suppl_1/D707 (ENSEMBL-2008)

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BaxevanisOuellette ch.1

• ncbi.nlm.nih.gov/RefSeq
• EMBL Genome Reviews
• GenPept http//www.ncbi.nlm.nih.gov/sites/entrez?
  dbprotein
• UniProt http//www.pir.uniprot.org/
• UniParc www.ebi.ac.uk/uniparc/
• IPI http//www.ebi.ac.uk/IPI/IPIhelp.html

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RefSeq

• January 11, 2008 RefSeq Release 27 available for
  FTP This release includes
• Proteins 4,426,609 Organisms 4,926 Available
  at ftp//ftp.ncbi.nih.gov/refseq/release/ To
  receive announcements of future RefSeq releases
  and incremental large updates please subscribe to
  NCBI's refseq-announce mail list refseq-announce

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More databases to know

• ExPASy, Expert Protein Analysis System
  http//www.expasy.org/
• SwissProt and TrEMBL at ExPASy
  http//www.expasy.org/sprot/
• UniProt/SwissProt by EBI www.ebi.ac.uk/swissprot
• UniProt PIR, protein information resource
  http//pir.georgetown.edu/
• www.uniprot.org ? beta.uniprot.org

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More NCBI DBs

• GenPept (compiled from SwissProt, PIR, PRF, PDB,
  and translations from annotated coding regions in
  GenBank and RefSeq http//www.ncbi.nlm.nih.gov/si
  tes/entrez?dbprotein
• The NCBI Entrez Taxonomy http//www.ncbi.nlm.nih.
  gov/sites/entrez?dbtaxonomy

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Sequence Formats

• ASCII TEXT
• There are at least a couple of dozen sequence
  formats in existence at the moment.
• Nucleotide (DNA or RNA) sequences are usually
  stored in the IUBMB standard codes. Similarly,
  protein sequences are usually stored in the IUPAC
  standard one-letter codes.

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IUPAC characters

• A adenine
• C cytosine
• G guanine
• T thymine
• U uracil
• R G A (puRine)
• Y T C (pYrimidine)
• K G T (keto)
• M A C (amino)
• S G C ltstrong interaction, 3H bondsgt
• W A T ltweak interaction, 2H bondsgt
• B G T C (not-A, follows B)
• D G A T (not C, follows C in the alphabet)
• H A C T (not G, follows G in the alphabet)
• V G C A (not-T (not-U), V follows U)
• N A G C T (any)

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Amino Acid Symbols
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Amino Acid Symbols -2
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Amino Acid Symbols -3
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Single-letter code recognition sequences for type
II restriction endonucleases
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FASTA format

• gtgi27827140gbBX094486.1BX094486 BX094486
  NCI_CGAP_Kid5 Homo sapiens cDNA clone
  IMAGp998D214660 IMAGE1901156 5', mRNA sequence
  AGGGCAAGGAGTAAAGGTGGCTGGGTGTGGGTCCGTTGAAGCGAGCCGCC
  TCCAGCCCTGTTGAACTGGTGGGCCCAGGGACTGGAGCGGGATTGAAAGG
  GATCTTGCTCTCCCTTGAAGCCTCGAGTTGCAGCGATTTCAGTGTCTTCT
  CTCCCTGTGTAAGCCTGTCTGGGTGTTTAGGCTGAACTACAGCCACCCCC
  TCTCCCGGGGGTGTGCAGGCCAGGGACTGGCCAGGCAGCCATGGCTGACG
  AGAAGACCTTCCGGATCGGCTTCATTGTGCTGGGGCTTTTCCTGCTGGCC
  CTCGGTACGTTCCTCATGAGCCATGATCGGCCCCAGGTCTACGGCACCTT
  CTATGCCATGGGCAGCGTCATGGTGATCGGGGGCATCATCTGGAGCATGT
  GCCAGTGCTACCCCAAGATCACCTTCGTCCCTGCTGACTCTGACTTTCAA
  GGCATCCTCTCCCCAAAGGCCATGGGCCTGCTGGAGAA

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FASTA format

• Starts from gt
• N omore than 80 characters in the header
• Sequence Codes
• A-Adenosine C-Cytidine G-Guanine T-Thymidine
  U-Uracil R-G or A (puRine) N - any
• http//en.wikipedia.org/wiki/Fasta_format

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Sample questions

• What records could be written in FASTA format?
  Genes, proteins, whole genomes, one or multiple
  sequences, aligned sequences, metabolic pathways?

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EMBL format

• A sequence file in EMBL format can contain
  several sequences. One sequence entry starts
  with an identifier line ("ID"), followed by
  further annotation lines. The start of the
  sequence is marked by a line starting with "SQ"
  and the end of the sequence is marked by two
  slashes ("//").
• ID AB000263 standard RNA PRI 368 BP.
• XX
• AC AB000263
• XX
• DE Homo sapiens mRNA for prepro cortistatin like
  peptide, complete cds.
• XX
• SQ Sequence 368 BP
• acaagatgcc attgtccccc ggcctcctgc tgctgctgct
  ctccggggcc acggccaccg 60
• ctgccctgcc cctggagggt ggccccaccg gccgagacag
  cgagcatatg caggaagcgg 120
• caggaataag gaaaagcagc ctcctgactt tcctcgcttg
  gtggtttgag tggacctccc 180
• aggccagtgc cgggcccctc ataggagagg aagctcggga
  ggtggccagg cggcaggaag 240
• gcgcaccccc ccagcaatcc gcgcgccggg acagaatgcc
  ctgcaggaac ttcttctgga 300
• agaccttctc ctcctgcaaa taaaacctca cccatgaatg
  ctcacgcaag tttaattaca 360
• gacctgaa 368 //

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GENBANK (GBK) format

• LOCUS AB000263 368 bp mRNA linear PRI 05-FEB-1999
• DEFINITION Homo sapiens mRNA for prepro
  cortistatin like peptide, complete cds.
• ACCESSION AB000263
• ORIGIN
• 1 acaagatgcc attgtccccc ggcctcctgc tgctgctgct
  ctccggggcc acggccaccg
• 61 ctgccctgcc cctggagggt ggccccaccg gccgagacag
  cgagcatatg caggaagcgg
• 121 caggaataag gaaaagcagc ctcctgactt tcctcgcttg
  gtggtttgag tggacctccc
• 181 aggccagtgc cgggcccctc ataggagagg aagctcggga
  ggtggccagg cggcaggaag
• 241 gcgcaccccc ccagcaatcc gcgcgccggg acagaatgcc
  ctgcaggaac ttcttctgga
• 301 agaccttctc ctcctgcaaa taaaacctca cccatgaatg
  ctcacgcaag tttaattaca
• 361 gacctgaa

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GENBANK (GBK) format

• ACCESSION BX094486
• VERSION BX094486.1 GI27827140
• source 1..488 /organism"Homo sapiens"
  /mol_type"mRNA" /db_xref"taxon9606"
  /clone"IMAGp998D214660 IMAGE1901156"
  /tissue_type"2 pooled tumors (clear cell type)"
  /lab_host"DH10B" /clone_lib"NCI_CGAP_Kid5"
  /note"Organ kidney Vector pT7T3D-PacI
  Site_1 Not I Site_2 Eco RI 1st strand cDNA
  was primed with a Not I - oligo(dT) primer 5'

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Compare again to FASTA format

• gtgi27827140gbBX094486.1BX094486 BX094486
  NCI_CGAP_Kid5 Homo sapiens cDNA clone
  IMAGp998D214660 IMAGE1901156 5', mRNA sequence
  AGGGCAAGGAGTAAAGGTGGCTGGGTGTGGGTCCGTTGAAGCGAGCCGCC
  TCCAGCCCTGTTGAACTGGTGGGCCCAGGGACTGGAGCGGGATTGAAAGG
  GATCTTGCTCTCCCTTGAAGCCTCGAGTTGCAGCGATTTCAGTGTCTTCT
  CTCCCTGTGTAAGCCTGTCTGGGTGTTTAGGCTGAACTACAGCCACCCCC
  TCTCCCGGGGGTGTGCAGGCCAGGGACTGGCCAGGCAGCCATGGCTGACG
  AGAAGACCTTCCGGATCGGCTTCATTGTGCTGGGGCTTTTCCTGCTGGCC
  CTCGGTACGTTCCTCATGAGCCATGATCGGCCCCAGGTCTACGGCACCTT
  CTATGCCATGGGCAGCGTCATGGTGATCGGGGGCATCATCTGGAGCATGT
  GCCAGTGCTACCCCAAGATCACCTTCGTCCCTGCTGACTCTGACTTTCAA
  GGCATCCTCTCCCCAAAGGCCATGGGCCTGCTGGAGAA

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ASN.1

• Abstract Syntax Notation One (ASN.1) is a formal
  language for abstractly describing messages to be
  exchanged among an extensive range of
  applications involving the Internet, intelligent
  network, cellular phones, ground-to-air
  communications, electronic commerce, secure
  electronic services, interactive television,
  intelligent transportation systems, Voice Over IP
  and others.
• Messages defined using ASN.1 can be encoded in
  XML format and visualized

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ASN.1

• Seq-entry seq id general db "dbEST" ,
  tag id 16732958 , embl accession "BX094486" ,
  version 1 , gi 27827140 ,
• title "BX094486 NCI_CGAP_Kid5 Homo sapiens cDNA
  clone IMAGp998D214660 IMAGE1901156 5'." ,
• subtype clone-lib , name "NCI_CGAP_Kid5" ,
  subtype tissue-type , name "2 pooled tumors
  (clear cell type)" ,
• orgname name binomial genus "Homo" , species
  "sapiens" , mod subtype other , subname
  "Organ kidney Vector pT7T3D-PacI Site_1 ,
  

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ASN.1

• Abstract Syntax Notation One (ASN.1) is a formal
  language for abstractly describing messages to be
  exchanged among an extensive range of
  applications involving the Internet, intelligent
  network, cellular phones, ground-to-air
  communications, electronic commerce, secure
  electronic services, interactive television,
  intelligent transportation systems, Voice Over IP
  and others. Due to its streamlined encoding
  rules, ASN.1 is also reliable and ideal for
  wireless broadband and other resource-constrained
  environments. Its extensibility facilitates
  communications between newer and older versions
  of applications. In a world of change, ASN.1 is
  core technology, constantly adapting to new
  technologies.ASN.1 is a critical part of our
  daily lives it's everywhere, but it works so
  well it's invisible
• The standardized XML Encoding Rules (XER) allow
  ASN.1 specifications (modules) to be used as
  ASN.1 schemas against which XML documents can be
  validated. As a result, the ASN.1 language now
  competes with other XML schema languages, but it
  has some additional benefits over them.

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ASN.1

• One of the difficulties in making use of data
  from a database is in interpreting the format of
  the data. A common approach in the past has been
  to release data in a so-called flat file format.
  However, this format fails to preserve the
  inherent relationships in a more complex data
  model--for example, a relational database
  management system such as the Sybase software
  used for the Plant Genome Database.
• A better approach is to make the data available
  according to a specification defined in a data
  description language. Abstract Syntax Notation 1
  (ASN.1) is one type of data description language.
  Data from the Plant Genome Database is to be made
  available in the ASN.1 format in addition to its
  primary means of access, which will be on-line.
  Also, data exchange between NAL and our
  collaborators will most likely occur using ASN.1.
  The article by Jim Ostell of the National Library
  of Medicine's National Center for Biotechnology
  Information describes ASN.1 and some advantages
  in its use
• http//www.nal.usda.gov/pgdic/Probe/v2n2/using.htm
  l

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SEQUENCE ANALYSIS TOOLS

• EMBOSS analysis tools
• http//www.molbiol.ox.ac.uk/analysis_tools/EMBOSS/
  index.shtml
• Pasteur Analysis tools
• http//bioweb2.pasteur.fr/intro-en.html
• ExPASy http//www.expasy.ch/tools/
• SEQOOL
• http//www.biossc.de/seqool/index.html

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Sequence Analysis Software

• Vector NTI
• Accelrys GCG
• Reda-Soft Textco
• BioSoftware, Inc.
• GCK
• Virtual Cloning SuiteTM

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• Vector NTI Advance 10
• Vector NTISequence analysis, annotation, and
  illustration restriction mapping recombinant
  molecule design, including Gateway and TOPO
  cloning in silico gel electrophoresis
  mol.bio-data management
• AlignXMultiple sequence alignment of proteins
  and DNAs
• ContigExpressDNA sequence assembly and
  sequencing project management using the CAP 3
  algorithm
• GenomBenchAnalysis and annotation of reference
  genomic DNA sequences
• BioAnnotatorFunctional annotation of DNAs and
  proteins

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MATLAB BIOINFORMATICS TOOLBOX DEMO
http//www.mathworks.com/access/helpdesk/help/tool
box/bioinfo/index.html?/access/helpdesk/help/toolb
ox/bioinfo/ug/fp6010dup1.htmlhttp//www.mathworks
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product_page.html