Extended Data Fig. 4: Targeted amplicon sequencing for evaluating endogenous targeted pseudouridylation.
From: Enhancing RNA base editing on mammalian transcripts with small nuclear RNAs
a, Schematic of BID-Seq to infer pseudouridylation of endogenous targeted mRNA by deletion rate. b, BID-Seq deletion rates on synthetic RNA standards (100% U and 100% Ψ at targeted base) for CFTR reporter locus. n = 1 technical replicate per condition. c, Effective pseudouridylation performance by transfection in HEK293T cells of guided U > Ψ snRNAs with (g)8 and (c)8 linkers versus H/ACA box snoRNA on CFTR reporter. Pseudouridylation difference significance versus snoRNA: ****, *****: p < 1e-4, 1e-5 (one-way ANOVA, Bonferroni correction for multiple comparisons) (snoRNA IDUA:CFTR 3e-9, CFTR snoRNA:(c)8 1e-5, CFTR snoRNA:(g)8 2e-6). Error bars reflect standard error of mean. n = 4 biological replicates per condition. d, Schematic of targeted amplicon CMC sequencing to infer pseudouridylation of endogenous targeted mRNA by mutation/deletion rate. e, Targeted amplicon CMC sequencing mutation/deletion rates on synthetic RNA standards (100% U and 100% Ψ at targeted base) for endogenous ACTB, EEF2, and RPS6 loci. n = 2 technical replicates per condition.
