Extended Data Fig. 9: Validation of Cas1-2/3 interactions with the repeat.
a, Time-course integration reactions to compare rate of integration into I-E and I-F repeats downstream of a I-F leader. b, Quantification of time-course experiments to determine the impact of 19 mutations associated with swapping the I-F repeat for the I-E repeat, on integration. Leader-side integration is indistinguishable. But spacer-side integration is slower into the I-E repeat. The mean and standard deviation of three replicate experiments are shown. c, Time-course integration reactions to measure the impact of I-F repeat mutations on integration rate. d, Time-course integration reactions to measure the impact of I-F repeat mutations on integration rate, in the context of a Cas1E184A mutation. The Cas1E184A mutation is expected to disrupt 5′ G recognition, but also impacts stability of the Cas1-2/3 complex (Extended Data Fig. 5). e, Quantification of time-course experiments to determine the impact of I-F repeat mutations on integration rate, in the context of WT Cas1-2/3. The mean and standard deviation of three replicate experiments are shown. f, Quantification of time-course experiments to determine the impact of I-F repeat mutations on integration rate, in the context of Cas1E184A-2/3. In panels e and f, no integration occurs into either the ‘G1A,T2A,A28C’ or ‘G1A,T2A’ repeat mutations, and the datapoints for these plots overlaps at roughly Y = 0 over the time course. The mean and standard deviation of three replicate experiments are shown. Each Urea-PAGE gel was run once.
