CRISPR-KRISPR: a method to identify on-target and random insertion of donor DNAs and their characterization in knock-in mice

Genome Biol. 2022 Oct 25;23(1):228. doi: 10.1186/s13059-022-02779-8.

Abstract

CRISPR tools can generate knockout and knock-in animal models easily, but the models can contain off-target genomic lesions or random insertions of donor DNAs. Simpler methods to identify off-target lesions and random insertions, using tail or earpiece DNA, are unavailable. We develop CRISPR-KRISPR (CRISPR-Knock-ins and Random Inserts Searching PRotocol), a method to identify both off-target lesions and random insertions. CRISPR-KRISPR uses as little as 3.4 μg of genomic DNA; thus, it can be easily incorporated as an additional step to genotype founder animals for further breeding.

Keywords: CIRCLE-seq; CRISPR; CRISPR-KRISPR; Donor DNA; Intronic region; Knock-in; Off target; Random insertion; Repeat element.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • CRISPR-Cas Systems*
  • Clustered Regularly Interspaced Short Palindromic Repeats*
  • DNA / genetics
  • Gene Editing / methods
  • Gene Knock-In Techniques
  • Genome
  • Mice

Substances

  • DNA