Detecting mutations from single DNA molecules is crucial in many fields but challenging. Next-generation sequencing (NGS) affords tremendous throughput but cannot directly sequence double-stranded DNA molecules (‘single duplexes’) to discern the true mutations on both strands. Here we present Concatenating Original Duplex for Error Correction (CODEC), which confers single duplex resolution to NGS. CODEC affords 1,000-fold higher accuracy than NGS, using up to 100-fold fewer reads than duplex sequencing. CODEC revealed mutation frequencies of 2.72 × 10⁻⁸ in sperm of a 39-year-old individual, and somatic mutations acquired with age in blood cells. CODEC detected genome-wide, clonal hematopoiesis mutations from single DNA molecules, single mutated duplexes from tumor genomes and liquid biopsies, microsatellite instability with 10-fold greater sensitivity and mutational signatures, and specific tumor mutations with up to 100-fold fewer reads. CODEC enables more precise genetic testing and reveals biologically significant mutations, which are commonly obscured by NGS errors.

检测单个 DNA 分子的突变在许多领域都至关重要，但也具有挑战性。新一代测序 (NGS) 提供巨大的通量，但无法直接对双链 DNA 分子（“单双链体”）进行测序以辨别两条链上的真正突变。在这里，我们提出了串联原始双工纠错 (CODEC)，它为 NGS 提供单双工分辨率。CODEC 的准确度比 NGS 高 1,000 倍，而使用的读数比双工测序少 100 倍。CODEC揭示了一名39岁个体精子中的突变频率为2.72 × 10 ^{-8 ，以及血细胞中随着年龄增长而获得的体细胞突变。}CODEC 可检测来自单个 DNA 分子的全基因组克隆造血突变、来自肿瘤基因组和液体活检的单个突变双链体、灵敏度和突变特征提高 10 倍的微卫星不稳定性，以及读数减少多达 100 倍的特定肿瘤突变。CODEC 能够实现更精确的基因检测，并揭示具有生物学意义的突变，而这些突变通常会被 NGS 错误所掩盖。