Fig. 1: Continuous prime editing in MMR-deficient cells at two endogenous loci produces near complete installation of intended edits.
From: A benchmarked, high-efficiency prime editing platform for multiplexed dropout screening
a, Western blot analysis of K562 cells (parental) and clonal derivatives stably expressing indicated prime editor protein from the AAVS1 safe-harbor locus, either with (PEmaxKO) or without (PE2, PEmax) genetic disruption of MLH1. Analysis after 1 month of culture post transduction with pegRNA constructs (from same cell populations as in b, d and e). b, Percentages of cells with expression of marker for prime editor construct (EGFP driven by IRES2 from the same transcript as the prime editor protein). Analysis over 1 month of culture post transduction with (e)pegRNA constructs, data and error bars represent mean ± s.d. of n = 4 independent biological samples for each cell line. c, Schematic of prime editing over time, with intended edit shown in cyan. d, Percentages of sequencing reads containing HEK3 +1 T>A (left) or DNMT1 +6 G>C (right) and no errors recovered from the indicated cells edited with either a pegRNA or epegRNA over 1 month. Edits are specified such that +1 and +6 represent nucleotide positions downstream from the Cas9(H840A) nick, with the +6 edit targeting within the PAM. Day 0 represents the timepoint at which cells were transduced with (e)pegRNA constructs. e, Percentages of sequencing reads containing either only the HEK3 +1 T>A (left) or DNMT1 +6 G>C (right) substitution (green) or errors (gray) from cells sampled 28 days post transduction of epegRNA constructs. Data and error bars in d and e represent mean ± s.d. (n = 3 independent biological replicates). c, Created with BioRender.com.
