Journal article

Detection of low-frequency DNA variants by targeted sequencing of the Watson and Crick strands

Joshua D Cohen, Christopher Douville, Jonathan C Dudley, Brian J Mog, Maria Popoli, Janine Ptak, Lisa Dobbyn, Natalie Silliman, Joy Schaefer, Jeanne Tie, Peter Gibbs, Cristian Tomasetti, Nickolas Papadopoulos, Kenneth W Kinzler, Bert Vogelstein

NATURE BIOTECHNOLOGY | NATURE RESEARCH | Published : 2021

Abstract

Identification and quantification of low-frequency mutations remain challenging despite improvements in the baseline error rate of next-generation sequencing technologies. Here, we describe a method, termed SaferSeqS, that addresses these challenges by (1) efficiently introducing identical molecular barcodes in the Watson and Crick strands of template molecules and (2) enriching target sequences with strand-specific PCR. The method achieves high sensitivity and specificity and detects variants at frequencies below 1 in 100,000 DNA template molecules with a background mutation rate of 100-fold.

Grants

Awarded by Medical Research Future Fund Investigator Grant


Awarded by National Institutes of Health


Funding Acknowledgements

We thank the individuals who participated in this study for their courage and generosity. We also thank M. Hoang, S. Sur, A. Mattox, A. Pearlman and members of the Ludwig Center at Johns Hopkins for insightful and helpful scientific discussions. We are grateful to C. Blair and K. Judge for expert technical and administrative assistance and to E. Cook for illustrative assistance. This work was supported by The Lustgarten Foundation for Pancreatic Cancer Research, The Marcus Foundation, The Virginia and D.K. Ludwig Fund for Cancer Research, The Conrad N. Hilton Foundation, The John Templeton Foundation, Medical Research Future Fund Investigator Grant (APP1194970) and National Institutes of Health grants (T32 GM007309, U01 CA230691-01, P50 CA228991, U01 CA200469, R37 CA230400-01, and U01 CA152753).