DNA Manipulation Here

I found this powerpoint online for what looks to be prepared for CDC and Homeland Security. Since it is in pdf format, I will link you to the baa.st.dhs.gov... meland+security+ft+collins&cd=12&hl=en&ct=clnk&gl=us&client=safari" target="_blank" class="postlink">Google Cache

What is being discussed is the genetic mutation of strains and also anthrax letters.

This is the html version of the file baa.st.dhs.gov...
Google automatically generates html versions of documents as we crawl the web.
Page 1
Forensic Analysis using SNP
and VNTR Signatures
Dr. Paul Keim
Northern Arizona University
And
Translational Genomics Research Institute (TGen)
Page 2
Biothreat Agents
• B. anthracis
• Y. pestis
• Francisella tularensis
• Burkholderia mallei, pseudomallei
• Brucella melitensis, abortus, etc.
• E. coli O157:H7
• Salmonella
Page 3
Molecular Microbial Forensics
• Molecular Assays
• Reference Databases
– Population studies
• Real-life Validation Studies
• Models and Theory
• Molecular Assays
• Reference Databases
– Population studies
• Real-life Validation Studies
• Models and Theory
Page 4
A genome is a
complex and
interesting
place to work.
Not all loci are
created equal
for diagnostics.
Genome Mining
Page 5
High Resolution “DNA Fingerprinting”
• Multiple-Locus VNTR Analysis (MLVA)
– High Discrimination markers – rapidly mutating.
• Single Nucleotide Polymorphisms (SNPs)
– Low Discrimination markers, but very stable!
• Multiple-Locus VNTR Analysis (MLVA)
– High Discrimination markers – rapidly mutating.
• Single Nucleotide Polymorphisms (SNPs)
– Low Discrimination markers, but very stable!
Page 6
Evolutionary Scale
and Mutation Rate
Evolutionary Scale
and Mutation Rate
Slow
Fast
10-3
10-4
10-5
10-6
10-7
10-8
10-9
10-10
Mutation Rate
SNR
VNTRs
SNR
VNTRs
Complex VNTRS
SNPs
Page 7
Tandemly Repeated
Sequences
Tandemly Repeated
Sequences
Genomic regions that vary greatly
among worldwide isolates.
VNTR
Page 8
Genomic Tandem Arrays
E. coli K12
B. anthracis Ames
Y. pestis CO92
Page 9
DNA Sequence Evolution
A
G
Transversion
Mutations
Transversion
Mutations
Kimura 2-parameter model
C
T
Transition
Mutations
Transition
Mutations
Page 10
Two Approaches to SNP Discovery:
• Limited Discovery:
•Multiple-Locus Sequence Typing
Appropriate when diversity is high.
• Exhaustive Discovery:
•Whole Genome Sequencing
Appropriate when diversity is low.
• Limited Discovery:
•Multiple-Locus Sequence Typing
Appropriate when diversity is high.
• Exhaustive Discovery:
•Whole Genome Sequencing
Appropriate when diversity is low.
Page 11
SNPs
Sequence-Based Differences
Page 12
Canonical Characters
• A phylogenetic character (such as a
SNP) that defines an evolutionary point.
• In practice, SNPs are chosen to
represent a whole groups of SNPs that
mark a common phylogenetic branch.
• A phylogenetic character (such as a
SNP) that defines an evolutionary point.
• In practice, SNPs are chosen to
represent a whole groups of SNPs that
mark a common phylogenetic branch.
Page 13
World Wide B. anthracis
Canonical SNPs
Marking major branches
and specific strains
Ames
KNP
French
C
NA-1
Page 14
Ames-Specific RT-PCR
b)
a)
Page 15
B. anthracis PHRANA (1067 isolates)
B. anthracis PHRANA (1067 isolates)
Canonical SNPs
MLVA-15
SNR-4
(208 GTs)
(476 GTs)
Page 16
Single Colony
Mutational Rate
Estimation
Mutational Rate
Estimation
10 Transfers X 96 lineages = 2.0 x 104 Generations
10 Transfers X 96 lineages = 2.0 x 104 Generations
Page 17
Mutation Rate Models
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1
0
50000
100000
150000
200000
250000
300000
350000
Generations Separating Two Isolates
Probability
0
1
2
3
4
5
Page 18
Anthrax Letter Attacks
Page 19
Page 20
• 15 VNTR Analysis
• 269 Isolates
• 47 genotypes
57
NA-1
136
NA-1
North
America
Anthrax
North
America
Anthrax
Matt Van Ert
unpublished
AI.a
Page 21
Molecular Forensics Analysis
Testimony in Court
Molecular Forensics Analysis
Testimony in Court
• “A match – Inclusion”
– Failure to exclude, could be the source.
• “Exclusion”
– Very different and unlikely the same.
• “Kind of Close?”
– Maybe yes? Maybe no?
• “A match – Inclusion”
– Failure to exclude, could be the source.
• “Exclusion”
– Very different and unlikely the same.
• “Kind of Close?”
– Maybe yes? Maybe no?
Page 22
Collaborators
• TIGR
– Jacques Ravell, Tim Read
• CDC Atlanta
• CDC Atlanta
– Alex Hoffmaster, Tanja Popovic
• CDC Ft. Collins
– May Chu, Ken Gage, Jennifer Lowell
• Louisiana State University
– Martin Hugh-Jones. Pamala Coker
• Institut Pasteur
– Agnes Fouet, Michele Mock, Elisabeth Carniel
• Umeå Sweden Group
• Umeå Sweden Grou
– Anders Johansson, Mats Forsman, Anders
Sjöstedt
• Livermore National Lab
– Gary Andersen, Emilio Garcia, Patrick Chain,
Peter Agron
• Los Alamos
– Paul Jackson, Rich Okinaka, Karen Hill
• TIGR
– Jacques Ravell, Tim Read
– Alex Hoffmaster, Tanja Popovic
• CDC Ft. Collins
– May Chu, Ken Gage, Jennifer Lowell
• Louisiana State University
– Martin Hugh-Jones. Pamala Coker
• Institut Pasteur
– Agnes Fouet, Michele Mock, Elisabeth Carniel
p
– Anders Johansson, Mats Forsman, Anders
Sjöstedt
• Livermore National Lab
– Gary Andersen, Emilio Garcia, Patrick Chain,
Peter Agron
• Los Alamos
– Paul Jackson, Rich Okina
• Keim Lab Characters
– Talima Pearson, Joseph Busch,
– Tatum Simonson, Shane Rhoton
– Jana U’Ren, Ryan Easterday,
– Matt Van Ert, Meghan Dukerich,
– Lynn Huynh, Shaylan Zanecki,
– Zack Jay, Michelle Maldonado,
– Amy Vogler, Jessica Girard,
– Kimothy Smith, Leo Kenefic,
Ryan Easterday, Chris Allender
– Jason Farlow, Dave Wagner,
– Christine Keys, Lance Price, Dan
Solomon, Sasha Klevytska, Jim
Schupp
• Keim Lab Characters
– Talima Pearson, Joseph Busch,
– Tatum Simonson, Shane Rhoton
– Jana U’Ren, Ryan Easterday,
– Matt Van Ert, Meghan Dukerich,
– Lynn Huynh, Shaylan Zanecki,
– Zack Jay, Michelle Maldonado,
– Amy Vogler, Jessica Girard,
– Kimothy Smith, Leo Kenefic,
Ryan Easterday, Chris Allender
– Jason Farlow, Dave Wagner,
– Christine Keys, Lance Price, Dan
Solomon, Sasha Klevytska, Jim
Schupp
ka, Karen Hill
Page 23
Research Funding
Dept. of Homeland Security
Bio-Chem Countermeasures
Department of Justice
Federal Bureau of Investigation
National Institutes of Health
Pathogen Evolution -NIGMS

hmmm.

What you guys think?

[edit on 6-10-2009 by 12.21.12]

[edit on 6-10-2009 by 12.21.12]

I am having trouble posting the link so here is the link to the pdf document.

baa.st.dhs.gov...

Heres a little more information

www.cdc.gov...

Single Nucleotide Polymorphism Typing of Bacillus anthracis from Sverdlovsk Tissue

The 1979 accidental release of Bacillus anthracis spores in the northeastern quadrant of Sverdlovsk (today's Ekaterinburg) in the former Soviet Union resulted in a relatively large-scale incident of inhalation anthrax (1). Histologic examinations of formalin-fixed, paraffin-embedded tissue samples from patients affected by this incident showed pathologic changes (2). Paraffin-embedded material from 11 of these patients was also subjected to molecular DNA analysis to demonstrate the presence of B. anthracis plasmid markers (3). These tissue samples appeared to contain 3–4 allelic variants of B. anthracis, based on a single variable-number tandem repeat (VNTR) marker (vrrA), which suggests that the material contained multiple strains of this species (4).

An early global comparison of the entire pagA gene sequence from 26 diverse B. anthracis isolates and a single nucleotide polymorphism (SNP)–rich region (307 bp) of the samples from Sverdlovsk found 7 SNPs in this gene (5). Two of these SNPs were unique to 3 of the Sverdlovsk samples. However, the distribution of the remaining 5 SNPs separated the 26 diverse isolates and the remaining Sverdlovsk samples into clusters that were consistent with diversity groups previously described by amplified fragment–length polymorphism (AFLP) analysis of a larger subset of isolates (6). The AFLP analysis had separated a collection of 78 isolates into 5 diversity groups. Although only 307 bp of the Sverdlovsk tissue samples were sequenced, a single SNP (pagA SNP 3602) placed 7 of 10 Sverdlovsk tissue samples into a large diversity group called western North America (WNA) (5).

uhm 12.21.12,

This may seem oftopic but i know you posted alot on the thread:
Omega codes and new bible story of the truth from Brandon Levon.

I posted a question here:

www.abovetopsecret.com...

See last post of mine: posted on 4-10-2009 @ 08:36 PM

Could you help me? I'm sorry if its offtopic but i cant post a new thread since i'm new and have to post 20 posts.