Precision gene editing

Francisella novicida Cas9 (FnCas9) has low cellular editing ability which limits its therapeutic utilities. Here, the authors rationally engineer the protein to develop enhanced FnCas9 (enFnCas9) variants with high on-target editing efficiency, high precision, broadened target range, and flexible base editing outcomes.

Site-directed insertion is a powerful approach for generating mutant lines but, historically, technically challenging. Here, the authors demonstrate CRIMP, an optimised protocol and universal toolkit (CRIMPkit), to greatly improve the efficacy of generating and studying mutant lines.

Engineered sex ratio distorters have been proposed as a powerful component of genetic control strategies designed to suppress harmful insect pests. Here the authors show that sex ratio distorters behave in unexpected ways in malaria mosquitoes, offering new paths for genetic pest control by targeting sex ratios.

Cas9 remains bound to DNA after cleavage and its removal is required for DNA double-strand break repair. Here, the authors show that the HLTF translocase disrupts the Cas9- DNA post-cleavage complexes in a process that requires the HLTF HIRAN domain and ATPase activity.

OliTag-seq, a specific and reproducible in-cellulo assay for CRISPR/Cas9 off-target analysis, can improve site cleavage efficiency and the identification of off-target sites.

CRISPR-Cas enzymes are increasingly entering clinical trials for gene therapies but can sometimes alter DNA off-target. We present an approach to modify the guide RNA to effectively eliminate off-target activity for target sequences of interest.

A survey of known human DNA editing sites with an RNA editing site prediction algorithm suggests APOBEC-mediated RNA editing may produce some of the same protein variants, with the possibility of affecting multiple areas of health.