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Fig. 7 | Molecular Cancer

Fig. 7

From: Strategies to overcome the main challenges of the use of CRISPR/Cas9 as a replacement for cancer therapy

Fig. 7

Nickase systems consisting of one or two nickases. H840 and D10 are two amino acids found in the Cas9 endonuclease protein that are involved in the cutting of one DNA strand by the enzyme. The RuvC domain contains the amino acid H840, while the HNH domain has the amino acid D10. The non-targeted strand is cleaved by Cas9 H840A, while the gRNA-targeting strand is cleaved by Cas9 D10A. Cas9 can only cut the strand complementary to the gRNA in a single nickase; however, a pair of sgRNA-Cas9n complexes can nick both strands at once (paired nickases). Additional concerns for gRNA design when using paired nickases include creating a 5’ overhang, the spacing between the two gRNAs, and the relative position of the two gRNA target sites

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