High-fidelity endonuclease variant HypaCas9 facilitates accurate allele-specific gene modification in mouse zygotes

Commun Biol. 2019 Oct 10:2:371. doi: 10.1038/s42003-019-0627-8. eCollection 2019.

Abstract

CRISPR/Cas9 has been widely used for the efficient generation of genetically modified animals; however, this system could have unexpected off-target effects. In the present study, we confirmed the validity of a high-fidelity Cas9 variant, HypaCas9, for accurate genome editing in mouse zygotes. HypaCas9 efficiently modified the target locus while minimizing off-target effects even in a single-nucleotide mismatched sequence. Furthermore, by applying HypaCas9 to the discrimination of SNP in hybrid strain-derived zygotes, we accomplished allele-specific gene modifications and successfully generated mice with a monoallelic mutation in an essential gene. These results suggest that the improved accuracy of HypaCas9 facilitates the generation of genetically modified animals.

Keywords: CRISPR-Cas9 genome editing; Genetic engineering.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CRISPR-Associated Protein 9* / genetics
  • CRISPR-Associated Protein 9* / metabolism
  • CRISPR-Cas Systems* / physiology
  • Female
  • Gene Editing / methods*
  • Male
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Mutation
  • Polymorphism, Single Nucleotide
  • Zygote* / metabolism

Substances

  • CRISPR-Associated Protein 9