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Sequencing small genomic targets with high efficiency and extreme accuracy


The detection of minority variants in mixed samples requires methods for enrichment and accurate sequencing of small genomic intervals. We describe an efficient approach based on sequential rounds of hybridization with biotinylated oligonucleotides that enables more than 1-million-fold enrichment of genomic regions of interest. In conjunction with error-correcting double-stranded molecular tags, our approach enables the quantification of mutations in individual DNA molecules.

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Figure 1: High on-target recovery with sequential rounds of capture.
Figure 2: Removal of sequencing artifacts by duplex sequencing.

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Research reported in this publication was supported by the US National Institutes of Health under award numbers NCI P01-CA77852, R01-CA160674 and R33-CA181771 to L.A.L. and NCI U10-CA180861, P01-CA018029, R01-CA175008 and R01-CA175215 to J.P.R. We thank T. Walsh and M. Lee for assistance with DNA sequencing.

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M.W.S., E.J.F., M.J.P., K.S.R.-B., L.D.T., J.P.R. and L.A.L. contributed to experimental design. M.W.S., E.J.F. and M.J.P. performed the experiments in the paper and analyzed data. E.J.F., L.D.T. and J.P.R. contributed patient samples. M.W.S. and L.A.L. wrote the manuscript.

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Correspondence to Lawrence A Loeb.

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M.W.S. and L.A.L. are named on a patent application regarding duplex sequencing that has been filed by the University of Washington.

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Schmitt, M., Fox, E., Prindle, M. et al. Sequencing small genomic targets with high efficiency and extreme accuracy. Nat Methods 12, 423–425 (2015).

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