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Fig. 2 | Genome Biology

Fig. 2

From: Expansion of the prime editing modality with Cas9 from Francisella novicida

Fig. 2

Expansion of the targetable range of prime editing by using FnCas9(H969A)-RT. a Schematics of the target gene (c-Myc) recognized by SpCas9(H840A)-RT and FnCas9(H969A)-RT. b Direct comparison of the efficiency (%) of base insertion by SpCas9(H840A)-RT (Top) and FnCas9(H969A)-RT (Bottom). c NGS result of site-specific base insertion induced by SpCas9(H840A)-RT and FnCas9(H969A)-RT. Each position and efficiency (%) in which the inserted AA bases are indicated to the left and right of the target sequence, respectively. d Engineered residues (Glu 1369, Glu 1449, Arg 1556) in FnCas9 for YG PAM recognition (PDB: 5B2O). e Target sequence for YG PAM recognition in the c-Myc gene commonly targeted using SpCas9(H840A)-RT, FnCas9(H969A)-RT and engineered FnCas9(H969A, E1369R, E1449H, R1556A)-RT. f The result of site-specific base insertion at YG PAM target induced by SpCas9(H840A)-RT and engineered FnCas9(H969A, E1369R, E1449H, R1556A)-RT using PE3 form delivery. g Comparison of the efficiency (%) of base insertion at YG(Y = C, T) PAM target by using SpCas9(H840A)-RT and engineered FnCas9(H969A, E1369R, E1449H, R1556A)-RT according to (f). P values are calculated using a two-way ANOVA, Dunnett test (ns: not significant, *P = 0.0332, **P = 0.0021, ***P = 0.0002, ****P < 0.0001). h Phi-chart demonstration for pathogenic SNP coverage of SpCas9(H840A)-RT or FnCas9(H969A)-RT prime editing. The nicking point counted as (N) bp upstream from the PAM (NGG or YG) is indicated at the bottom of the phi chart. Covered ratio (%) = targetable SNP number/total SNP number ×100, uncovered ratio (%) = 100 − covered ratio (%). Each histogram was plotted by applying standard error of the mean values to repeated experimental values (n = 3)

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