Accurate whole human genome sequencing using reversible terminator chemistry

David R. Bentley, Shankar Balasubramanian, Harold P. Swerdlow, Geoffrey P. Smith, John Milton, Clive G. Brown, Kevin P. Hall, Dirk J. Evers, Colin L. Barnes, Helen R. Bignell, Jonathan M. Boutell, Jason Bryant, Richard J. Carter, R. Keira Cheetham, Anthony J. Cox, Darren J. Ellis, Michael R. Flatbush, Niall A. Gormley, Sean J. Humphray, Leslie J. IrvingMirian S. Karbelashvili, Scott M. Kirk, Heng Li, Xiaohai Liu, Klaus S. Maisinger, Lisa J. Murray, Bojan Obradovic, Tobias Ost, Michael L. Parkinson, Mark R. Pratt, Isabelle M.J. Rasolonjatovo, Mark T. Reed, Roberto Rigatti, Chiara Rodighiero, Mark T. Ross, Andrea Sabot, Subramanian V. Sankar, Aylwyn Scally, Gary P. Schroth, Mark E. Smith, Vincent P. Smith, Anastassia Spiridou, Peta E. Torrance, Svilen S. Tzonev, Eric H. Vermaas, Klaudia Walter, Xiaolin Wu, Lu Zhang, Mohammed D. Alam, Carole Anastasi, Ify C. Aniebo, David M.D. Bailey, Iain R. Bancarz, Saibal Banerjee, Selena G. Barbour, Primo A. Baybayan, Vincent A. Benoit, Kevin F. Benson, Claire Bevis, Phillip J. Black, Asha Boodhun, Joe S. Brennan, John A. Bridgham, Rob C. Brown, Andrew A. Brown, Dale H. Buermann, Abass A. Bundu, James C. Burrows, Nigel P. Carter, Nestor Castillo, Maria Chiara E. Catenazzi, Simon Chang, R. Neil Cooley, Natasha R. Crake, Olubunmi O. Dada, Konstantinos D. Diakoumakos, Belen Dominguez-Fernandez, David J. Earnshaw, Ugonna C. Egbujor, David W. Elmore, Sergey S. Etchin, Mark R. Ewan, Milan Fedurco, Louise J. Fraser, Karin V. Fuentes Fajardo, W. Scott Furey, David George, Kimberley J. Gietzen, Colin P. Goddard, George S. Golda, Philip A. Granieri, David E. Green, David L. Gustafson, Nancy F. Hansen, Kevin Harnish, Christian D. Haudenschild, Narinder I. Heyer, Matthew M. Hims, Johnny T. Ho, Adrian M. Horgan, Katya Hoschler, Steve Hurwitz, Denis V. Ivanov, Maria Q. Johnson, Terena James, T. A. Huw Jones, Gyoung Dong Kang, Tzvetana H. Kerelska, Alan D. Kersey, Irina Khrebtukova, Alex P. Kindwall, Zoya Kingsbury, Paula I. Kokko-Gonzales, Anil Kumar, Marc A. Laurent, Cynthia T. Lawley, Sarah E. Lee, Xavier Lee, Arnold K. Liao, Jennifer A. Loch, Mitch Lok, Shujun Luo, Radhika M. Mammen, John W. Martin, Patrick G. McCauley, Paul McNitt, Parul Mehta, Keith W. Moon, Joe W. Mullens, Taksina Newington, Zemin Ning, Bee Ling Ng, Sonia M. Novo, Michael J. O'Neill, Mark A. Osborne, Andrew Osnowski, Omead Ostadan, Lambros L. Paraschos, Lea Pickering, Andrew C. Pike, Alger C. Pike, D. Chris Pinkard, Daniel P. Pliskin, Joe Podhasky, Victor J. Quijano, Come Raczy, Vicki H. Rae, Stephen R. Rawlings, Ana Chiva Rodriguez, Phyllida M. Roe, John Rogers, Maria C. Rogert Bacigalupo, Nikolai Romanov, Anthony Romieu, Rithy K. Roth, Natalie J. Rourke, Silke T. Ruediger, Eli Rusman, Raquel M. Sanches-Kuiper, Martin R. Schenker, Josefina M. Seoane, Richard J. Shaw, Mitch K. Shiver, Steven W. Short, Ning L. Sizto, Johannes P. Sluis, Melanie A. Smith, Jean Ernest Sohna Sohna, Eric J. Spence, Kim Stevens, Neil Sutton, Lukasz Szajkowski, Carolyn L. Tregidgo, Gerardo Turcatti, Stephanie Vandevondele, Yuli Verhovsky, Selene M. Virk, Suzanne Wakelin, Gregory C. Walcott, Jingwen Wang, Graham J. Worsley, Juying Yan, Ling Yau, Mike Zuerlein, Jane Rogers, James C. Mullikin, Matthew E. Hurles, Nick J. McCooke, John S. West, Frank L. Oaks, Peter L. Lundberg, David Klenerman, Richard Durbin, Anthony J. Smith

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2332 Citations (Scopus)

Abstract

DNA sequence information underpins genetic research, enabling discoveries of important biological or medical benefit. Sequencing projects have traditionally used long (400-800 base pair) reads, but the existence of reference sequences for the human and many other genomes makes it possible to develop new, fast approaches to re-sequencing, whereby shorter reads are compared to a reference to identify intraspecies genetic variation. Here we report an approach that generates several billion bases of accurate nucleotide sequence per experiment at low cost. Single molecules of DNA are attached to a flat surface, amplified in situ and used as templates for synthetic sequencing with fluorescent reversible terminator deoxyribonucleotides. Images of the surface are analysed to generate high-quality sequence. We demonstrate application of this approach to human genome sequencing on flow-sorted X chromosomes and then scale the approach to determine the genome sequence of a male Yoruba from Ibadan, Nigeria. We build an accurate consensus sequence from >30× average depth of paired 35-base reads. We characterize four million single-nucleotide polymorphisms and four hundred thousand structural variants, many of which were previously unknown. Our approach is effective for accurate, rapid and economical whole-genome re-sequencing and many other biomedical applications.

Original languageEnglish
Pages (from-to)53-59
Number of pages7
JournalNature
Volume456
Issue number7218
DOIs
Publication statusPublished - 6 Nov 2008
Externally publishedYes

Bibliographical note

Funding Information:
Acknowledgements The authors acknowledge the advice of A. Williamson, T. Rink, S. Benkovic, J. Berriman, J. Todd, R. Waterston, S. Eletr, W. Jack, M. Cooper, T. Brown, C. Reece and R. Cook during this work; E. Margulies for assistance with data analysis; M. Shumway for assistance with data submission; and the contributions of the administrative and support staff at all the institutions. This research was supported in part by The Wellcome Trust (to H.L., A.Sc., K.W., N.P.C, B.N.L., J.R., M.E.H. and R.D.), the Biotechnology and Biological Sciences Research Council (BBSRC) (to S.B. and D.K.), the BBSRC Applied Genomics LINK Programme (to A.Sp. and C.L.B.) and the Intramural Research Program of the National Human Genome Research Institute, National Institutes of Health (to N.F.H. and J.C.M.). S. Balasubramanian and D. Klenerman are inventors and founders of Solexa Ltd.

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