Fig. 2
From: Engineering the next-generation of CAR T-cells with CRISPR-Cas9 gene editing
CRISPR/Cas9 components and Cas9 enzyme class variants. (Left) Endonuclease Cas9 can cleave a specific site within DNA as determined by the complementarity of the synthetic (crispr) crRNA to the target DNA strand, together with a (trans-activating) tracrRNA hairpin to provide structural stability. The crRNA and tracrRNA make up a single guide RNA (sgRNA) complex. Additional specificity is provided by the protospacer adjacent motif (PAM) that directs Cas9 to cut 3 base pairs upstream. a Cas9 cuts DNA via its HNH and RuvC domains that each cut a single DNA strand, resulting in a double-stranded break. b Mutations in one domain or both domains restrict(s) Cas9 to perform single-strand nicking (nCas9) or (c) to possess no catalytic activity (dCas9). Created with BioRender.com