Fig. 2: PRINS editing with modified pegRNAs in human cell lines.
From: Modified pegRNAs mitigate scaffold-derived prime editing by-products
a PRINS editing at PCSK9, CTLA4, HBEGF and AAVS1 genomic loci in four different human cell lines (K562, HEK293T, HeLa and HepG2) using electroporation of PEn mRNA in combination with synthetic pegRNA, rSp-pegRNA or C3-pegRNA to install indicated small insertions (ins.). Editing outcomes were analyzed by amplicon-seq and quantified using CRISPResso2 in the prime editing mode. Plots show mean ± SD of n = 3 biological replicates. Prime edits precise = intended prime edits; Prime edits all (no scaffold) = precise prime edits + prime edits co-occurring with indels not derived from scaffold; Scaffold incorporated = prime edits with at least one additional nucleotide derived from scaffold; Indels = non-prime edited insertions or deletions. b Quantification of data from (a). “Precision score” was calculated as total number of amplicon-seq reads with precise prime edit per overall editing. “Prime edits containing scaffold” was calculated as total number of amplicon-seq prime edited reads with scaffold integration per total prime edited reads. Error bars represent mean ± SD. Datapoints represent 4 different cell line experiments in (a). Statistical difference was determined using Student’s t test (paired, two-tailed). *P < 0.05, **P < 0.01, ***P < 0.001. Calculated P values: PCSK9: pegRNA vs. rSp-pegRNA = 0.0032, pegRNA vs. C3-pegRNA = 0.0014; CTLA4: pegRNA vs. rSp-pegRNA = 0.0039, pegRNA vs. C3-pegRNA = 0.0057; HBEGF: pegRNA vs. rSp-pegRNA = 0.0034, AAVS1: pegRNA vs. rSp-pegRNA = 0.0154. Source data are provided as a Source Data file.
