Use of paired Cas9-NG nickase and truncated sgRNAs for single-nucleotide microbial genome editing.

ABSTRACT: The paired nickases approach, which utilizes clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated proteins (Cas) nickase and dual guide RNA, has the advantage of reducing off-target effects by being able to double the target sequence. In this study, our research utilized the Cas9-NG nickase variant to minimize PAM sequence constraints, enabling the generation of paired nicks at desired genomic loci. We performed a systematic investigation into the formation sites for double nicks and the design of donor DNA within a bacterial model system. Although we successfully identified the conditions necessary for the effective formation of double nicks in vivo, achieving single-nucleotide level editing directly at the target sites in the genome proved challengi

SUBMITTER: Jeong SH 

PROVIDER: S-EPMC11464485 | biostudies-literature | 2024

REPOSITORIES: biostudies-literature