Extended Data Fig. 3: Supercoiled DNA with canonical and spaced PAMs were nicked using single-nuclease SpCas9 mutants.
From: Shifted PAMs generate DNA overhangs and enhance SpCas9 post-catalytic complex dissociation
(a) HNH-SpCas9 (D10A) nickase (containing an active HNH domain but inactive RuvC domain) was assembled with various dgRNAs into RNPs to incise the corresponding target double-stranded circular DNA containing either a canonical PAM or a shifted PAM. (b) As with the prior panel but using RuvC-SpCas9 (H840A) nickase (containing an active RuvC domain but inactive HNH domain). (c) Efficiencies of SSB formation are quantified, normalized and shown as blue diamonds for HNH-SpCas9(D10A) and red dots for RuvC-SpCas9(H840A). Error bars indicate SEM (independent experiments, n = 2).
