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 >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.

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

新发现朝1000美元/人的宏伟基因组测序计划迈近了坚实一步

图片说明：新方法测序无需进行DNA扩增。

（图片来源：Getty）