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Article
Nature Methods - 4, 931 - 936 (2007)
Published online: 14 October 2007; Corrected online: 21 October 2007 | doi:10.1038/nmeth1110

Multiplex amplification of large sets of human exons

Gregory J Porreca1, 8, Kun Zhang1, 7, 8, Jin Billy Li1, Bin Xie2, Derek Austin2, Sara L Vassallo1, Emily M LeProust3, Bill J Peck3, Christopher J Emig4, Fredrik Dahl5, 7, Yuan Gao2, 6, George M Church1, 8 & Jay Shendure1, 7, 8

1  Department of Genetics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, Massachusetts 02115, USA.

2  Center for the Study of Biological Complexity, Virginia Commonwealth University, 1000 W. Cary St. Richmond, Virginia 23284, USA.

3  Genomics Solution Unit, Agilent Technologies Inc., 5301 Stevens Creek Blvd., Santa Clara, California 95051, USA.

4  Codon Devices Inc., One Kendall Square, Building 300, Third Floor, Cambridge, Massachusetts 02139, USA.

5  Stanford Genome Technology Center, Clark Center W300, 318 Campus Drive, Stanford, California 94305, USA.

6  Department of Computer Science, Virginia Commonwealth University, 601 West Main Street, Richmond, Virginia 23284, USA.

7  Present addresses: Department of Bioengineering, University of California at San Diego, 9500 Gilman Dr., La Jolla, California 92093, USA (K.Z.), Complete Genomics Inc., 2071 Stierlin Court, Suite 100, Mountain View, California 94043, USA (F.D.), and Department of Genome Sciences, University of Washington, 1705 NE Pacific St., Seattle, Washington 98195, USA (J.S.).

8  These authors contributed equally to this work.

Correspondence should be addressed to Jay Shendure shendure@u.washington.edu or George M Church http://arep.med.harvard.edu/gmc/email.html

A new generation of technologies is poised to reduce DNA sequencing costs by several orders of magnitude. But our ability to fully leverage the power of these technologies is crippled by the absence of suitable 'front-end' methods for isolating complex subsets of a mammalian genome at a scale that matches the throughput at which these platforms will routinely operate. We show that targeting oligonucleotides released from programmable microarrays can be used to capture and amplify approx10,000 human exons in a single multiplex reaction. Additionally, we show integration of this protocol with ultra-high-throughput sequencing for targeted variation discovery. Although the multiplex capture reaction is highly specific, we found that nonuniform capture is a key issue that will need to be resolved by additional optimization. We anticipate that highly multiplexed methods for targeted amplification will enable the comprehensive resequencing of human exons at a fraction of the cost of whole-genome resequencing.
NOTE: In the version of this article initially published online,the affiliation for Jay Shendure was listed as Department of Computer Science, Virginia Commonwealth University,601 West Main Street,Richmond,Virginia 23284,USA. The correct affiliation should be Department of Genome Sciences,University of Washington,1705 NE Pacific St.,Seattle,Washington 98195,USA. The error has been corrected for all versions of the article.

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Nature Methods
ISSN: 1548-7091
EISSN: 1548-7105
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