title: Single-molecule DNA sequencing of a viral genome.
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.
authors.affiliation: Helicos BioSciences Corporation, One Kendall Square, Cambridge, MA 02139, USA. tharris@helicosbio.com
authors.forename: Timothy D
authors.initials: TD
authors.lastname: Harris
authors.forename: Phillip R
authors.initials: PR
authors.lastname: Buzby
authors.forename: Hazen
authors.initials: H
authors.lastname: Babcock
authors.forename: Eric
authors.initials: E
authors.lastname: Beer
authors.forename: Jayson
authors.initials: J
authors.lastname: Bowers
authors.forename: Ido
authors.initials: I
authors.lastname: Braslavsky
authors.forename: Marie
authors.initials: M
authors.lastname: Causey
authors.forename: Jennifer
authors.initials: J
authors.lastname: Colonell
authors.forename: James
authors.initials: J
authors.lastname: Dimeo
authors.forename: J William
authors.initials: JW
authors.lastname: Efcavitch
authors.forename: Eldar
authors.initials: E
authors.lastname: Giladi
authors.forename: Jaime
authors.initials: J
authors.lastname: Gill
authors.forename: John
authors.initials: J
authors.lastname: Healy
authors.forename: Mirna
authors.initials: M
authors.lastname: Jarosz
authors.forename: Dan
authors.initials: D
authors.lastname: Lapen
authors.forename: Keith
authors.initials: K
authors.lastname: Moulton
authors.forename: Stephen R
authors.initials: SR
authors.lastname: Quake
authors.forename: Kathleen
authors.initials: K
authors.lastname: Steinmann
authors.forename: Edward
authors.initials: E
authors.lastname: Thayer
authors.forename: Anastasia
authors.initials: A
authors.lastname: Tyurina
authors.forename: Rebecca
authors.initials: R
authors.lastname: Ward
authors.forename: Howard
authors.initials: H
authors.lastname: Weiss
authors.forename: Zheng
authors.initials: Z
authors.lastname: Xie
mesh_terms: D000465:Algorithms; D017104:Bacteriophage M13 / Q000235:genetics*; D019295:Computational Biology / Q000379:methods; D017931:DNA Primers; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics*; D016679:Genome, Viral; D009154:Mutation; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA / Q000379:methods*; D012984:Software; D013698:Templates, Genetic
chemical_list: D017931:DNA Primers; D004279:DNA, Viral