Fig. 2: Characterization of SuperFi and its variants in mammalian cells.

a–d (a, b) EGFP disruption activity on 8 target sites and c, d indels measured on 6 endogenous sites with a, c 20G-sgRNAs and the same sites targeted with b, d 21G-sgRNAs, normalized to their 20G-sgRNA counterparts, presented on a scatter dot plot. Data are related to Supplementary Fig. 5b, c. e On-target activity of various SpCas9 variants as measured in EGFP disruption assay on 40 target sites presented on a scatter dot plot. Data are related to Supplementary Fig. 6. f Normalized EGFP disruption activity of SpCas9 nucleases with perfectly matching (n = 18) and partially mismatching (n = 162) 20G-sgRNAs. Dots are shown for each variant with each mismatching spacer position, provided that the on-target activity exceeded 70% with every SpCas9 variant; data are omitted otherwise. a–f The median and interquartile ranges are shown; data points are plotted as open circles representing the mean of triplicates. Summary of target and primer sequences, EGFP disruption and NGS data are reported in Supplementary Data 1–3. Statistical significance was assessed by RM one-way ANOVA, statistical details (only statistically significant differences compared to SuperFi are shown) and p-values are available in Methods and in Supplementary Data 6 (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001).