Extended Data Fig. 5: iBE has low DNA off target activity.
From: Generation of precision preclinical cancer models using regulated in vivo base editing
a. Schematic of experimental set up in mouse embryonic stem cells (ESCs). mESCs containing iBE knock in were transduced with LRT2B-gRNA vector and selected for gRNA expression. sgRNA+ cells were plated with and without dox for 6 days after which cells were plated at low density for clonal outgrowth without dox. 3 pools of 10 clones were picked for each dox conditions across to gRNA targeted cell lines (sgRNAs = Apc.Q1405X and Pik3ca.E545K). In total, 12 pools of 10 clones were sequenced at 800-1000-fold coverage across the MSK-IMPACT cancer gene set. b. Pie chart display of frequency of C>T or C>other SNVs found in pooled clones for each condition (on and off dox) for both sgRNAs. c. Sequencing analysis at cancer gene sites in cell conditions (right) described in a. Solid blue boxes represent on-target activity of the sgRNA, dotted orange boxes signify on-target ‘bystander’ editing within the gRNA window. d. Quantification of C>T and C>other SNVs found across both targets. 2-way ANOVA test for multiple comparisons was used to evaluate statistical significance across conditions. Data are presented as mean values ± s.e.m. p-values are displayed.
