Cytosine base editor generates substantial off-target single-nucleotide variants in mouse embryos

TLDR

Unintended genomic modifications from gene editing limit therapeutic use, and current off‑target detection methods generally fail in vivo or to identify single‑nucleotide changes. The authors employed DNA‑repair‑protein recruitment to CRISPR‑Cas9 breaks, single‑blastomere injections in two‑cell mouse embryos, and whole‑genome sequencing of edited rice plants to unbiasedly detect off‑target edits. New in‑vivo monitoring methods were reported, and cytosine base editors were found to induce numerous single‑nucleotide variants in mouse and rice, whereas adenine editors did not. References: Wienert et al.; Zuo et al.; Jin et al.; Science, this issue, pp.

Abstract

Spotting off-targets from gene editing Unintended genomic modifications limit the potential therapeutic use of gene-editing tools. Available methods to find off-targets generally do not work in vivo or detect single-nucleotide changes. Three papers in this issue report new methods for monitoring gene-editing tools in vivo (see the Perspective by Kempton and Qi). Wienert et al. followed the recruitment of a DNA repair protein to DNA breaks induced by CRISPR-Cas9, enabling unbiased detection of off-target editing in cellular and animal models. Zuo et al. identified off-targets without the interference of natural genetic heterogeneity by injecting base editors into one blastomere of a two-cell mouse embryo and leaving the other genetically identical blastomere unedited. Jin et al. performed whole-genome sequencing on individual, genome-edited rice plants to identify unintended mutations. Cytosine, but not adenine, base editors induced numerous single-nucleotide variants in both mouse and rice. Science , this issue p. 286 , p. 289 , p. 292 ; see also p. 234