Gt(ROSA)26Sor^{em1(CAG-cas9*,-EGFP)Dvs}

Sequences include CMV enhancer/chicken beta-actin promoter (CAG), a loxP-STOP(3xSV40 polyA)-loxP cassette (LSL), a 3xFLAG sequence, a mammalian codon-optimized cas9 gene (derived from Streptococcus pyogenes CRISPR associated protein 9 [SpCas9]), a P2A ribosomal skip cleavage peptide sequence, an enhanced green fluorescent protein gene (EGFP), a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) and a bovine growth hormone polyA sequence (BGHpA). To facilitate insertion into the Gt(ROSA)26Sor locus on chromosome 6 via homology directed repair (HDR), the LSL-Cas9-P2A-EGFP sequences were co-injected with a large piece of Gt(ROSA)26Sor locus homologous DNA. (J:101977)