a, Effect of changing the sequence surrounding the target C on editing efficiency in vitro. The deamination yield of 80% of targeted strands (40% of total sequencing reads from both strands) for C₇ in the protospacer sequence 5′-TTATTT(C₇)GTGGATTTATTTA-3′ was defined as 1.0, and the relative deamination efficiencies of substrates containing all possible single-base mutations at positions 1–6 and 8–13 are shown. b, Positional effect of each NC motif on editing efficiency in vitro. Each NC target motif was varied from positions 1 to 8 within the protospacer as indicated in the sequences shown on the right. The PAM is shown in blue. The graph shows the percentage of total DNA sequencing reads containing T at each of the numbered target C positions following incubation with BE1. Note that the maximum possible deamination yield in vitro is 50% of total sequencing reads (100% of targeted strands). Values and error bars reflect the mean and standard deviation of three (a) or two (b) independent biological replicates performed on different days.