Artificial sgRNAs engineered for genome editing with new Cas12b orthologs

a Graphical overview of the eight Cas12b orthologs tested in this study. Sizes (amino acids) are indicated. b T7EI assay results indicating the genome targeting activities of the eight Cas12b orthologs directed by their cognate sgRNAs in human 293T cells. Red triangles indicate the cleaved bands. c, d T7EI assay results indicating the genome targeting activities of the eight Cas12b orthologs directed by AasgRNA (c) and AksgRNA (d) in human 293T cells. Red triangles indicate the cleaved bands. e Maps of bacterial genomic loci corresponding to DiCas12b and TcCas12b. The two Cas12b loci have no CRISPR array. f, g T7EI assay results indicating the genome targeting activities of AaCas12b, DiCas12b, and TcCas12b directed by AasgRNA (f) and AksgRNA (g) in human 293T cells. Red triangles indicate the cleaved bands. h T7EI assay results indicating the simultaneous multiplex genome targeting mediated by TcCas12b combined with AksgRNAs in human 293T cells. i Schematic illustration of the secondary structures of artificial sgRNA scaffold 13 (artsgRNA13). j T7EI assay results indicating the simultaneous multiplex genome targeting mediated by TcCas12b combined with artsgRNA13s in human 293T cells. k T7EI analysis of average targeted mutation efficiencies induced at five different endogenous loci by AaCas12b, TcCas12b, AsCas12a, and FnCas12a. n？=？