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

Research output: Contribution to journalArticlepeer-review

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 <5 × 10–7 mutants per base pair (bp). We demonstrate that it can evaluate mutations in a single amplicon or simultaneously in multiple amplicons, assess limited quantities of cell-free DNA with high recovery of both strands and reduce the error rate of existing PCR-based molecular barcoding approaches by >100-fold.

Original languageEnglish (US)
JournalNature biotechnology
DOIs
StateAccepted/In press - 2021

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Molecular Medicine
  • Biomedical Engineering

Fingerprint

Dive into the research topics of 'Detection of low-frequency DNA variants by targeted sequencing of the Watson and Crick strands'. Together they form a unique fingerprint.

Cite this