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Cas9

Difference between Cas9n (H840A) and Cas9n (N863A)?

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Mercedes Board Aug 29, 2022

Hi, I thought the most commonly used Cas9 nickases were D10A and H840A and I’m preparing to construct these two mutants by PCR, but then I came across an article that utilized D10A and N863A, so I’m wondering about the difference between H840A and N863A, which one should I use?
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Sudeshna Nandi Answered Aug 30, 2022

University of Innsbruck

Hi there, both H840A and N863A mutants inactivated the HNH domain of Cas9 protein, and a research (Nishimasu et al., 2014) claimed that H840A nickase has a reduced activity compared with N863A nickase. When using only one nickase in the experiment, the N863A nickase can be replaced with the D10A in all cases. And it is recommended to prepare these plasmids as an endotoxin-free maxiprep.

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