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Science 4 April 2008: Vol. 320. no. 5872, pp. 106 - 109 DOI: 10.1126/science.1150427
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Reports
Single-Molecule DNA Sequencing of a Viral Genome
Timothy D. Harris,1*
Phillip R. Buzby,1
Hazen Babcock,1
Eric Beer,1
Jayson Bowers,1
Ido Braslavsky,2
Marie Causey,1
Jennifer Colonell,1
James DiMeo,1
J. William Efcavitch,1
Eldar Giladi,1
Jaime Gill,1
John Healy,1
Mirna Jarosz,1
Dan Lapen,1
Keith Moulton,1
Stephen R. Quake,3
Kathleen Steinmann,1
Edward Thayer,1
Anastasia Tyurina,1
Rebecca Ward,1
Howard Weiss,1
Zheng Xie1
The full promise of human genomics will be realized only when the genomes of thousands of individuals can be sequenced for comparative analysis. A reference sequence enables the use of short read length. We report an amplification-free method for determining the nucleotide sequence of more than 280,000 individual DNA molecules simultaneously. A DNA polymerase adds labeled nucleotides to surface-immobilized primer-template duplexes in stepwise fashion, and the asynchronous growth of individual DNA molecules was monitored by fluorescence imaging. Read lengths of >25 bases and equivalent phred software program quality scores approaching 30 were achieved. We used this method to sequence the M13 virus to an average depth of >150 x and with 100% coverage; thus, we resequenced the M13 genome with high-sensitivity mutation detection. This demonstrates a strategy for high-throughput low-cost resequencing.
1 Helicos BioSciences Corporation, One Kendall Square, Cambridge, MA 02139, USA.
2 Department of Physics and Astronomy, Ohio University, Athens, OH 45701, USA.
3 Department of Bioengineering, Stanford University, and Howard Hughes Medical Institute, Stanford, CA 94305, USA.
* To whom correspondence should be addressed. E-mail: tharris{at}helicosbio.com
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