Single nucleotide substitutions effectively block Cas9 and allow for scarless genome editing in Caenorhabditis elegans.

ABSTRACT: In Caenorhabditis elegans, germline injection of Cas9 complexes is reliably used to achieve genome editing through homology-directed repair of Cas9-generated DNA breaks. To prevent Cas9 from targeting repaired DNA, additional blocking mutations are often incorporated into homologous repair templates. Cas9 can be blocked either by mutating the PAM sequence that is essential for Cas9 activity or by mutating the guide sequence that targets Cas9 to a specific genomic location. However, it is unclear how many nucleotides within the guide sequence should be mutated, since Cas9 can recognize "off-target" sequences that are imperfectly paired to its guide. In this study, we examined whether single-nucleotide substitutions within the guide sequence are sufficient to block Cas9 and allow for efficie

SUBMITTER: Medley JC 

PROVIDER: S-EPMC8733430 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature