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. Author manuscript; available in PMC: 2020 Apr 18.
Published in final edited form as: Mol Cell. 2019 Mar 5;74(2):296–309.e7. doi: 10.1016/j.molcel.2019.01.038

Figure 7. AcrIIC1/2/3 target hybrid SpyCas9 proteins.

Figure 7.

(A) Schematic of the SpyCas9NmeHNH-mediated DNA cleavage. The hybrid Cas9 enzyme, sgRNA, target strand, non-target strand, and PAM sequence are colored in gray, black, blue, yellow, and red, respectively. The RuvC and HNH domains of NmeCas9 cleave the non-target and target strands (black triangles), respectively.

(B) End-labelled DNA cleavage assays conducted using the hybrid SpyCas9NmeHNH protein to assess the cleavage on the target and non-target strands. The lanes for a given condition correspond to 1 and 2 h of reaction. Red stars depict fluorescein labels placed on the ends of DNA strands.

(C) DNA cleavage assays conducted using SpyCas9NmeHNH or NmeCas9 with or without AcrIIC3.

(D) DNA cleavage assays conducted using SpyCas9NmeHNH with or without AcrIIC1.

(E) End-labelled DNA cleavage assays conducted using the hybrid. The lanes for a given condition correspond to 1 and 2 h of reaction.

(F) DNA cleavage assays conducted using WT SpyCas9 or SpyCas9NmeBH with or without AcrIIC2.

See also Figure S7.