Homo sapiens and zebrafish Targeted Locus (Loci)

We find that pre-crRNAs comprising a full-length direct repeat (full-DR-crRNA) sequence with specific stem-loop G-C base substitutions exhibit increased editing efficiencies compared with the standard mature crRNA framework in cultured mammalian cell lines. In addition, we demonstrate in zebrafish embryos that the improved LbCpf1 and FnCpf1 nucleases in combination with these modified crRNAs display high mutagenesis efficiencies (more than 80% editing efficiency on a majority of tested targets) when delivered as ribonucleoprotein complexes. These results define a set of Cpf1 components with broad utility in vertebrate systems.