Bioinformatics%20at%20Promega%20Corporation

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Bioinformatics at Promega Corporation
Intro to Bioinformatics Biotec November 28,
2006 Ethan Strauss Sr. Scientist RD
Bioinformatics, Promega, Ethan.strauss_at_promega.c
om http//q7.com/ethan/molbio
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My Background

• Bachelors degree in biology
• PhD and work experience in Molecular Biology
• Eight years in Promega Technical Services
• Almost two years in Bioinformatics (officially)
• No formal computer training
• No formal bioinformatics training

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Bioinformatics at Promega Corporation

• Bioinformatics did not exists as a separate
  function until 2001
• One person 2001- 2005
• Two people 2005 - ?
• Bioinformatics supports primarily RD (100
  scientists)
• Mentor and train RD scientists
• Provide expertise for projects (120 requests per
  year)
• Propose and evaluate new acquisitions
• Liaison to IT department
• Manage bioinformatics infrastructure (15 tools)
• Develop new tools and adapt existing tools in
  house

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Bioinformatics Projects

• Programming
• Tools for internal and external Promega customers
• Plexor Primer Design System (https//www.promega
  .com/techserv/tools/plexor/logon.aspx)
• Biomath (http//www.promega.com/biomath/)
• siRNA Designer (http//www.promega.com/siRNADesi
  gner/)
• Sequence analysis for Excel and Microsoft
  Word(http//www.promega.com/enotes/features/fe002
  5.htm)
• Analysis of BLAST results
• Automated data retrieval (Web services)
• Database for tracking vector construction
• Database for keeping track of plasmid features

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Bioinformatics Projects

• Biocomputing (use of computers in biological
  research)
• Database searches
• data mining
• discovery research
• Primer design
• Blast analysis and interpretation
• Etc

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NCBI

• I recently took the Powerscripting course from
  NCBI
• NCBI has a lot of very powerful tools and
  databases.
• They are not as well documented as they might be.
  
• Check them out periodically.
• Databases at NCBI I was not aware of, but am now.
  
• Pub Med CentralArticles with free text
• 3D domain, structure, 3D structural information.
• GEO (Gene Expression Omnibus)Micorarray
  expression data
• There are many more which I see on the drop down
  list, but dont really know any thing about

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NCBI ftp site

• Most NCBI data is available by FTP from
  http//www.ncbi.nlm.nih.gov/Ftp/
• I have used it for a number of projects including
  an analysis of amino acid residue distribution
  for the first 11 positions of human and E. coli

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NCBI - Entrez Programming Utilities
Programatic access to Entrez http//eutils.ncbi.nl
m.nih.gov/entrez/query/static/eutils_help.html
Allows incorporation of entrez functionality
into third party tools http//www.promega.com/tech
serv/tools/plexor/NewQpcrProject.aspx Allows
automation of Entrez searchesAnalysis of large
datasetsAutomation of searches and
queries Accessable using HTTP or SOAP
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NCBI - Entrez Programming Utilities

• Programs available
• ESearch Searches and retrieves primary IDs and
  term translations and optionally retains results
  for future use in the user's environment.
• ESummary Retrieves document summaries from a
  list of primary IDs or from the user's
  environment.
• EFetch Retrieves records in the requested
  format from a list of one or more primary IDs or
  from the user's environment.
• ELink Checks for links from the query ID
  numbers to other Entrez databases
• EInfo Provides field index term counts, last
  update, and available links for each database.
• EPost Posts a file containing a list of primary
  IDs for future use in the user's environment to
  use with subsequent search strategies.

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NCBI - Entrez Programming Utilities
Lets try it! Go to http//www.ncbi.nlm.nih.gov/Cla
ss/wheeler/eutils/eu.html and play Now
try http//www.ncbi.nlm.nih.gov/Class/wheeler/euti
ls/epipe.html
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NCBI - Entrez Programming Utilities
These sorts of utilities can be access
programtically using Perl. See Demonstration
Programs at http//eutils.ncbi.nlm.nih.gov/entrez
/query/static/eutils_help.html
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NCBI - Entrez Programming Utilities
my utils "http//www.ncbi.nlm.nih.gov/entrez/eu
tils" my db ask_user("Database",
"Pubmed") my query ask_user("Query",
"zanzibar") my report ask_user("Report",
"abstract") my esearch "utils/esearch.fcgi?
dbdbretmax1usehistoryyterm" my
esearch_result get(esearch . query) print
"\nESEARCH RESULT esearch_result\n" esearch_re
sult mltCountgt(\d)lt/Countgt.ltQueryKeygt(\d)lt
/QueryKeygt.ltWebEnvgt(\S)lt/WebEnvgts my Count
1 my QueryKey 2 my WebEnv
3 print "Count Count QueryKey QueryKey
WebEnv WebEnv\n" my retstart my
retmax3 for(retstart 0 retstart lt Count
retstart retmax) my efetch
"utils/efetch.fcgi?rettypereportretmodetextr
etstartretstartretmaxretmax" .
"dbdbquery_keyQueryKeyWebEnvWebEnv"
print "\nEF_QUERYefetch\n" my
efetch_result get(efetch) print
"---------\nEFETCH RESULT(". (retstart
1) . . (retstart retmax) . ") ".
"efetch_result\n-----PRESS ENTER!!!-------\n"

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Bioinformatics Advice

• Be aware of bias in databases!
• Search Genbank (nucleotide) for HumanOrganism
  apoptosis. How many hits?
• Now try OrcinusOrganism apoptosisHow many
  hits?
• Can you conclude that Orcinus does not have
  apoptosis?

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Bioinformatics Advice

• Bioinformatics is changing and advancing very
  rapidly.
• Dont forget to notice what is new.
• NCBI now has 20 different databases. They had
  two only 3-5 years ago
• If you want to do something that you know cant
  be done, check again in two weeks!
• My standard computer can process the entire human
  genome for Restriction sites, ORFs etc in a few
  hours. Not long ago, the best computers couldnt
  even hold that much data!
• If old tools work, dont feel you need to use the
  newest tools.
• I still do much of my analysis with Microsoft
  Word

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LIMS Laboratory Information Management System
Goal Manage in-house DNA sequences and
associated data Eval UW-Madison Center for
Eukaryotic Structural Genomics Sesame
http//www.sesame.wisc.edu/ Sesame is designed
to organize and record data relevant to complex
scientific projects, to launch computer-controlled
processes, and to help decide about subsequent
steps on the basis of information available. The
Sesame system is based on the multi-tier
paradigm, and it consists of a framework and
application modules that carry out specific
tasks.Users interact with Sesame through a
series of web-based Java applet-applications
designed to organize data. It allows
collaborators on a given project to enter,
process, view, and extract relevant data,
regardless of location, so long as web access is
available. Data reside in an Oracle relational
database. Sesame serves as a digital laboratory
notebook and allows users to attach numerous
files and images
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Programming

• Tools for Promega customers
• Biomath (http//www.promega.com/biomath/)
• Basic calculations (Most can be done easily by
  hand)
• Simple code (Javascript)
• Established theory.
• Universal (not Promega specific)
• siRNA Designer(http//www.promega.com/siRNADesigne
  r/ )
• Complex calculations
• More complex code (VBScript)
• Rapidly evolving theory
• Partially Promega specific

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Programming

• Tools for Promega customers
• Plexor Primer Design (https//www.promega.com/tech
  serv/tools/plexor)
• Complex calculations
• Complex code (C.Net)
• Separate user interface and main calculations
• Multiple interacting modules
• Database integration
• Integration with Genbank (through a web service)
• Proprietary improvements on established theory
• Very Promega specific

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Programming

• Tools for internal use
• BLAST analysis of Plexor Primers
• Primer specificity is important
• BLAST can determine specificity, but output is
  very complex.
• Simplify
• Combine all hits from the same Gene
• Only show hits which could mis-prime
• Groups hits by species
• Allow sorting by species

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Programming

• Tools for internal use
• BLAST analysis of Plexor Primers

Initial BLAST results (1 page out of 30)
Analyzed BLAST results (complete!)
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Programming

• Tools for internal use
• Vector/Insert Database
• Promegas Flexi vector system has a very
  structured cloning procedure.
• RD has been making many different Flexi vector
  backbones with many inserts.
• Keeping track has been a problem.
• A database is in development

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Programming

• Tools for internal use

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Programming

• Internal Projects
• Which Restriction enzyme cuts least frequently in
  human ORFs?
• Method
• Download human Refseq database (ftp//ftp.ncbi.nih
  .gov/refseq/H_sapiens/)
• Load into local database
• Scan each sequence for each RE site
• The scan took 2-3 hours to complete

http//www.promega.com/pnotes/89/12416_11/12416_11
.pdf
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Programming

• Internal Projects
• Which human genes in Genbank are the most
  popular?
• Method
• Download Gene database (ftp//ftp.ncbi.nlm.nih.g
  ov/gene/)
• Download Gene Ontology information
  (http//www.geneontology.org/)
• Use web services to get pathway information from
  KEGG (http//www.genome.jp/kegg/)
• Use web services to get citation information from
  Pubmed (http//www.ncbi.nlm.nih.gov/entrez/query.f
  cgi?dbPubMed)
• Load all into local database
• Rank genes by desired criteria
• Size
• Function
• Localization
• Pathways
• Publications

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Database searches and data mining
Question Can you reformat this sequence for
me?Tool ReadSeq http//bimas.dcrt.nih.gov/molb
io/readseq Macros Question How many viral
proteins start with MetHis?Tool Hits database
motif searches http//hits.isb-sib.ch/ Question
How many different bacterial two-domain
proteins are known?Tool SCOP database
http//scop.berkeley.edu/ Question How do I
design PCR primers selective for bacterial
species X?Tool Ribosomal database 16s rRNA
alignment http//rdp.cme.msu.edu