Fig. 4: Dual inhibition approach in p53 mutant hPSCs.
From: Enhancing genome editing in hPSCs through dual inhibition of DNA damage response and repair pathways
A Sequences in 175R (Arginine) region of WT hESCs and TP53 mutant iPSCs (SES8). The amino acid of mutated SES8 was altered to H (Histidine) by one-base substitution. B Colony area calculated after Cas9 and PE gene editing in TP53 mutant iPSCs (SES8). Colony area calculated by ImageJ after gene editing in SES8. C C to T substitution ratio of each indicated target by AncBE4max plasmid with non-targeting siRNA and pcDNA 3.0 vector (Mock), siRNA targeting UNG (siUNG), p53DD expression vector (p53DD), and both siUNG and p53DD (BE4DI) at 4 genomic sites in SES8 (n = 3 except the designated replicates). D, E Normalized C to T (D) and C to G substitution (E) of Fig. 4C (n = 12 except the designated replicates). The C to T and C to G substitution ratio is normalized to Mock efficiency. F, G C to T conversion ratio (F) and total relative C to T and C to G (G) of AncBE4max and AncBEstem in SES8 (n = 3 except the designated replicates). H Relative prime editing efficiency of PE2 and PE4 (n = 4). I, J Prime editing efficiency of PE4max (PE4), PE4max with p53DD (PE4 + DD), PE5max (PE5) and PE5max with p53DD (PE5 + DD) (I) and PE4max, PE4stem, PE5max, and PE4stem (J) of indicated target (n = 3 except the designated replicates). K PE2 effects on the enrichment of TP53 mutant population. WT hPSCs and TP53 mutant hPSCs were mixed in a certain ratio, and then PE2 and pegRNA vectors were treated for gene editing. After 3 days, gDNA for each sample was harvested and the ratio of R175 mutation was tested by NGS. n always represents the biologically independent samples if not else described. Bars represent mean values, and error bars represent the S.D. of independent biological replicates. Detailed information on statistical analysis is listed in the “Statistical analysis” section. The source data of B–K are provided in the Source Data file.
