FIGURE 5.

The mechanisms of CRISPR/Cas9 system and the influence factors of off‐targets. CRISPR/Cas9 system is composed of Cas9 and a gRNA. Cas9‐gRNA complex identifies the strand including a PAM (NGG) at the 3′ end adjacent to the 20‐base pair target site. Cas9 consists of a α‐helical recognition (REC) lobe and nuclease lobe containing RuvC and HNH domain, which are joined by a bridge helix and cleave the two DNA strands simultaneously after conformation. The off‐target activities induced by CRISPR/Cas9 are affected by the non‐canonical PAM at off‐target sites, DNA accessibility, the structure of gRNA and Cas9 as well as the delivery methods of the Cas9‐gRNA compound.