Single-molecule DNA sequencing of a viral genome - PubMed (original) (raw)

. 2008 Apr 4;320(5872):106-9.

doi: 10.1126/science.1150427.

Phillip R Buzby, Hazen Babcock, Eric Beer, Jayson Bowers, Ido Braslavsky, Marie Causey, Jennifer Colonell, James Dimeo, J William Efcavitch, Eldar Giladi, Jaime Gill, John Healy, Mirna Jarosz, Dan Lapen, Keith Moulton, Stephen R Quake, Kathleen Steinmann, Edward Thayer, Anastasia Tyurina, Rebecca Ward, Howard Weiss, Zheng Xie

Affiliations

Single-molecule DNA sequencing of a viral genome

Timothy D Harris et al. Science. 2008.

Abstract

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 >150x 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.

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