Massively Parallel Profiling of RNA-targeting CRISPR-Cas13d.

ABSTRACT: Type VI CRISPR enzymes cleave target RNAs and are widely used for gene regulation, RNA tracking, and diagnostics. However, a systematic understanding of their RNA binding specificity and cleavage activation is lacking. Here, we describe RNA chip-hybridized association-mapping platform (RNA-CHAMP), a massively parallel platform that repurposes next-generation DNA sequencing chips to measure the binding affinity for over 10,000 RNA targets containing structural perturbations, mismatches, insertions, and deletions relative to the CRISPR RNA (crRNA). Deep profiling of Cas13d, a compact and widely used RNA nuclease, reveals that it does not require a protospacer flanking sequence (PFS) but is exquisitely sensitive to secondary structure within the target RNA. Cas13d binding is strongly penalize

SUBMITTER: Kuo HC 

PROVIDER: S-EPMC10081190 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature