Evaluation of Cas13d as a tool for genetic interaction mapping.

ABSTRACT: Mapping genetic interactions (GIs) is crucial for understanding genetic network complexity. In this study, we investigate the utility of Cas13d, a CRISPR system targeting RNA, for GI mapping and compare it to Cas9 and Cas12a, two DNA nucleases commonly used for GI mapping. We find that Cas13d induces faster target gene perturbation and generates more uniform cell populations with double perturbations than Cas9 or Cas12a. We then encounter Cas13d gRNA-gRNA interference when concatenating gRNAs targeting different genes into one gRNA array, which we overcome by a dual promoter gRNA expression strategy. Moreover, by concatenating three gRNAs targeting the same gene into one array, we are able to maximize the Cas13d-mediated knockdown effects. Combining these strategies enhances proliferation

SUBMITTER: El Kassem G 

PROVIDER: S-EPMC11828948 | biostudies-literature | 2025 Feb

REPOSITORIES: biostudies-literature