Fig. 4: IS607 TnpB-mediated genome editing in human cells.

a Scheme of the human cell line (HEK293F) genome-editing experiment. b The indel efficiencies of ISFba1 TnpB on endogenous loci EMX1-T1, EMX1-T2, VEGFA-T1, DNMT1-T1, DNMT1-T2 and PITX1-T1 in HEK293F cells, determined by next-generation sequencing (NGS). Data represent means ± SD of three biological replicates. Two-tailed unpaired t-test: ***P < 0.001, **P < 0.01. c NGS statistic of indel efficiency of ISFba1 TnpB on the endogenous locus EMX1-T1. d The indel efficiencies mediated by TnpBs of ISFba1, ISRin, ISFba4, ISEre1, ISAre, ISClsp3, ISDfo4, ISAfa and ISHun on the endogenous loci. Data represent means ± SD of three biological replicates. Two-tailed unpaired t-test: ***P < 0.001, **P < 0.01, *P < 0.05. The detailed information of different IS607 TnpB proteins is provided in Supplementary information, Table S2. The guide sequences are provided in Supplementary information, Table S5. e Heatmap representation of the ISFba1 TnpB cleavage efficiency with 60 single-nucleotide-mutated guide RNAs for EMX1-T1 target site. The identities of single base pair substitutions are indicated on the left; original guide sequence is shown at the bottom and highlighted in the heatmap (gray squares). The cleavage efficiencies were monitored by high-throughput sequencing. Modification efficiencies (increasing from white to blue) are normalized to the original guide sequence. 1–20, mismatch position 1–20. f Heatmap of the single mismatch tolerance of ISFba1 TnpB on the EMX1-T1, EMX1-T2, VEGFA-T1 target sites and SpCas9 on the EMX1 target site. 1–20, mismatch position 1–20; WT, original guide sequence. Data shown are representative of three independent experiments. The guide sequences are provided in Supplementary information, Table S5.