Improved Chemistry and Software Provides Higher Accuracy Single
Molecule Reads and Longer Readlengths to
Assembly
has completed a de novo sequence assembly of the Escherichia
coli O104:H4 strain responsible for the recent outbreak in
using its Single Molecule Real Time (SMRT™) technology, and sequenced 11
related bacterial strains (including six previously unsequenced strains
of the same serotype) for comparative analyses. An international team of
scientific experts on E. coli collaborated on the rapid
sequencing project to provide more comprehensive information about the
origins of the strain that gave rise to the deadly outbreak. The data
were generated using an early version of chemistry and software in
development at Pacific Biosciences for the next major PacBio RS product
upgrade, planned for the fourth quarter of 2011.
The data provided to the public domain includes a complete assembly of
the German outbreak strain, alignment to assemblies from other outbreak
isolates, and sequences for 11 related Enteroaggregative E. coli
strains. The project demonstrates the ability to produce a PacBio-only de
novo assembly for a complex microbial pathogen, and the power of
rapid sequencing of multiple genomes with the PacBio RS to
elucidate the evolutionary history of a pathogenic microbe. A summary of
the project appears on the company’s website at https://blog.pacificbiosciences.com.
The Pacific Biosciences scientific team, led by Chief Scientific Officer
experts on E. coli and infectious diseases for this project. The
collaborators include:
-
Karen Angeliki Krogfelt, Ph.D., Professor, Head of Unit,
Gastrointestinal Infections, Statens Serum Institut (SSI),
Denmark -
Flemming Scheutz , Ph.D., Head of theWHO Collaborating Centre for on Escherichia and Klebsiella,Reference and Research
SSI,
Denmark
In the U.S.:
-
James P. Nataro , M.D., Ph.D., Professor and Chair, Pediatrics,University of Virginia School of Medicine -
David A. Rasko , Ph.D., Assistant Professor,University of Maryland ,School of Medicine
Institute for Genome Sciences andDepartment of Microbiology and Immunology
-
Nadia Boisen , Ph.D., Research Scientist,Department of Pediatrics ,University of Virginia School of Medicine -
Matthew K. Waldor , M.D., Ph.D., Professor of Medicine atHarvard , Brigham and Women’s Hospital, and HMMIMedical School
“Using samples provided by our collaborators, we rapidly sequenced each
strain using a standard PacBio RS protocol that took on average
less than eight hours from sample preparation to sequencing results,”
said Dr. Schadt. “The ability to sequence the outbreak strain with reads
averaging 2,900 base pairs and our longest reads at over 7,800 bases,
combined with our circular consensus sequencing to achieve high single
molecule accuracy with a mode accuracy distribution of 99.9%, enabled us
to complete a PacBio-only assembly without having to construct
specialized fosmid libraries, perform PCR off the ends of contigs, or
other such techniques that are required to get to similar assemblies
with second generation DNA sequencing technologies.”
Dr. Krogfelt commented: “These high quality data will provide scientists
with more information about the genomic features of this strain that
could provide new markers for predicting the higher degree of
pathogenicity we are seeing with this outbreak. A more comprehensive
evolutionary view of this pathogen may also help identify markers for
antibiotic drug resistance that could be used in the future should other
related strains emerge. The complexity of this case proves that
international collaborations and communications are important in the
achievement of detailed scientific information.”
The data are available for the bioinformatics community at the PacBio
developer’s network (DevNet) web site (www.pacbiodevnet.com),
where a suite of open source tools and other resources designed for SMRT
sequence data are available to analyze the information. The data have
also been submitted to the
(NCBI) SRA database.
While not involved with the current
Institute has been testing the new version of the sequencing enzyme in
development as part of Pacific Biosciences’ early access program. “We
are seeing significant increases in readlength, with high quality runs
producing reads with average 2,000 base readlengths or more,” said
Nusbaum, co-director of the Genome Sequencing and Analysis program at
the
possibility to trade off readlength to increase accuracy will be
important for bringing the PacBio RS to an increasing application
space. We have been impressed with the rapid progress PacBio has made in
improving their technology in a relatively short amount of time.”
collaboration employed an early version of our chemistry and software in
development for the next major upgrade of our product, we are taking
this opportunity to share some of the specifications that we expect to
reach for that planned Q4 upgrade. As demonstrated by the E. coli
project, we are already seeing a big step up from the specifications for
the first commercial release in April, and we expect to achieve even
more significant performance increases.” While not yet fully optimized,
the specifications are based on flexible parameters with the capability
to produce:
-
An average of 2,700 base pair reads, with 5% of the reads achieving
5,100+ base pairs
-
Up to 90 megabases of mappable data per SMRT Cell
-
1,350 base pair reads with 2X circular consensus sequencing single
molecule accuracy of 93%
For more information about Pacific Biosciences, please visit www.pacificbiosciences.com.
You can also follow the company on twitter www.twitter.com/pacbio.
About Pacific Biosciences
Pacific Biosciences’ mission is to transform the way humankind acquires,
processes and interprets data from living systems through the design,
development and commercialization of innovative tools for biological
research. The company has developed a novel approach to studying the
synthesis and regulation of DNA, RNA and proteins. Combining recent
advances in nanofabrication, biochemistry, molecular biology, surface
chemistry and optics, Pacific Biosciences has created a powerful
technology platform called single molecule, real-time, or SMRT™,
technology. SMRT technology enables real-time analysis of biomolecules
with single molecule resolution, which has the potential to transform
the understanding of biological systems by providing a window into these
systems that has not previously been open for scientific study.
Forward-Looking Statements
This press release contains forward-looking statements. Forward-looking
statements may contain words such as “believe,” “may,” “estimate,”
“anticipate,” “continue,” “intend,” “expect,” “plan,” the negative of
these terms, or other similar expressions, and include the assumptions
that underlie such statements. Such statements include, but are not
limited to, statements regarding the Company’s SMRT technology. These
statements are subject to known and unknown risks and uncertainties that
could cause actual results to differ materially from those expressed or
implied by such statements, including but not limited to risks discussed
from time to time in documents
has filed with the
risks identified under the section captioned “Risk Factors” in its
recently filed Quarterly Report on Form 10-Q. All forward-looking
statements are based on estimates, projections and assumptions as of the
date hereof. Pacific Biosciences undertakes no obligation to update any
forward-looking statements.
Media:
For Pacific Biosciences
Litchfield, 415-793-6468
nicole@bioscribe.com
or
Investors:
Pacific
Biosciences
ir@pacificbiosciences.com
Source: Pacific Biosciences
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