Figure 1. Targeted integration of pGTag vector cargo DNA into a 5’ coding exon.

Short homology arms complementary to the 5’ (green) and 3’ (blue) sequences of the genomic target site are cloned on the 5’ and 3’ sides of the vector cargo DNA. The short homology arm cargo cassette is flanked by two universal guide RNA UgRNA sites. CRISPR/Cas9 simultaneously targets double-strand breaks at the sgRNA genomic target site and at the UgRNA sites flanking the cargo on the plasmid donor. Exonuclease end resection liberates single-stranded DNA in the vector homology arms that is complementary to the resected strands on the 5’ and 3’ sides of the genomic double-strand break. The complementary sequences direct homology mediated end joining integration of the cargo DNA at the exon target site. PAM sequences are underlined, and small red arrows indicate Cas9 cut sites in the genome and vector.