....13, Hu et al. first restored catalytic residues D10 and H840. To ensure that the clones were capable ...

...spacing D10A: nick sites separated by 37-68 bp H840A: nick sites separated by 51-68 bp Use D10A for HDR...use SpCas9 nickases as examples for this post. The D10A mutation inactivates the RuvC domain, so this nickase...preferences using HEK293 cells. They started by comparing D10A and H840A total editing efficiency in PAM-in and...higher in the PAM-out configuration. In addition, D10A editing efficiency is consistently higher than that...H840A, especially with smaller nick distances. Using D10A, they subsequently showed that editing efficiency...gRNAs are too close (7-23 nt nick distance). Both D10A and H840A are potent editors, but their indel profiles...profiles vary. In the PAM-out configuration, D10A tends to generate small deletions, while H840A is biased...

.... The double nickase system is based on the Cas9 D10A nickase described in Figure 4 of the Cong, et. al...

...instead of Cas9, this plasmid expresses Cas9n (a D10A nickase mutant) to nick, rather than cleave DNA....

... deletions (indel) by the catalytically impaired D10A nickase from ABE7.10 can be a problem. Therefore...

... other than NGG (Table-1). Type Nickase Cas9 (D10A) Deaminase PAM requirements Reference CBE SpCas9...

...enzymatic residues has been converted to an alanine (D10A or H840A), changing its double-strand cut to a single-stranded...

...expressed them along with Cas9 in the T-cell line 2D10. PCR amplification and Sanger sequencing of pooled...

... Cas9 variants are available: wt humanized Cas9, D10A nickase mutant (Cas9n), dCas9 (transcriptional repression...