Cas12a crRNA engineering

Design and validation of sixteen types of structurally engineered Cas12a crRNAs targeting various immunologically relevant loci. We designed structural modifications in the loop region, ranging from engineered breaks (STAR-crRNAs) to large gaps (Gap-crRNAs), as well as nucleotide substitutions, enable gene-cutting in the presence of various Cas12a nucleases. Moreover, we observe similar insertion rates of short HDR templates using the engineered crRNAs compared to the wild-type crRNAs, further demonstrating that the introduced modifications in the loop region lead to comparable genome editing efficiencies.