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. Author manuscript; available in PMC: 2023 Nov 5.
Published in final edited form as: Nat Chem Biol. 2023 Jul 3;19(11):1384–1393. doi: 10.1038/s41589-023-01380-9

Fig. 4 |. Structure-guided engineering of the AsCas12f gRNA.

Fig. 4 |

a, Indel frequencies mediated by enAsCas12f with engineered AsCas12f gRNAs at HEXA and PDCD1 loci. The structures of engineered gRNAs are shown in Supplementary Fig. 14. b, Structure of sgRNA-v2. c, Time-course in vitro DNA cleavage using full-length sgRNA and sgRNA-v2. The assay was conducted using enAsCas12f at 37 °C. Data points were fitted to one-phase exponential association curves. Gel images are provided in Supplementary Fig. 16. d, Indel frequencies mediated by the full-length sgRNA and sgRNA-v2 in complex with enAsCas12f at denoted genomic loci in HEK293T cells. e, Box-and-whisker plot of indel frequencies mediated by the full-length sgRNA or sgRNA-v2 in complex with enAsCas12f. All data points (n = 8 target sites) were plotted, with the centerline showing the median and the whiskers showing the minimum to the maximum. The boundaries of the box indicate the first and third quantiles. P values were determined by two-tailed paired Student’s t-test. f, Relative abundance of full-length sgRNA and sgRNA-v2 targeting HEXA and PDCD1 loci in HEK293T cells. Two independent replicates were carried out in a, c, and d.