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Open Access Highly Accessed Method

Multiplex target capture with double-stranded DNA probes

Peidong Shen1, Wenyi Wang2, Aung-Kyaw Chi1, Yu Fan2, Ronald W Davis1 and Curt Scharfe1*

Author Affiliations

1 Stanford Genome Technology Center, Stanford University, Palo Alto, CA 94304, USA

2 Department of Bioinformatics and Computational Biology, UT MD Anderson Cancer Center, Houston, TX 77030, USA

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Genome Medicine 2013, 5:50  doi:10.1186/gm454

Published: 29 May 2013

Abstract

Target enrichment technologies utilize single-stranded oligonucleotide probes to capture candidate genomic regions from a DNA sample before sequencing. We describe target capture using double-stranded probes, which consist of single-stranded, complementary long padlock probes (cLPPs), each selectively capturing one strand of a genomic target through circularization. Using two probes per target increases sensitivity for variant detection and cLPPs are easily produced by PCR at low cost. Additionally, we introduce an approach for generating capture libraries with uniformly randomized template orientations. This facilitates bidirectional sequencing of both the sense and antisense template strands during one paired-end read, which maximizes target coverage.