Figure 4.

E. coli genome-wide landscape of resistance to CRISPR/Cas9-induced lethal DNA DSBs. (A) Distribution of median sgRNA activity among all sgRNAs within each gene (for each of the three conditions, Cas9, eSpCas9 and Cas9 (ΔrecA)). The median sgRNA activity of negative control sgRNAs is zero because of the normalization step in the data processing (see Materials and Methods). (B) Genes with significant resistance to CRISPR/Cas9-induced DSBs were extracted ((median sgRNA activity ≥ 0 or (median sgRNA activity ≥ −1 and FPR ≥ 0.01)) and (≥5 sgRNAs)), giving rise to 192, 188 and 70 genes in the Cas9, eSpCas9 and Cas9 (ΔrecA) datasets, respectively. Shown is a distribution of deviations in the activities of sgRNAs targeting individual genes of these three sets. The experimental noise as quantified by negative control sgRNA deviations is shown as dotted lines. (C) Median activity among all sgRNAs belonging to each gene is plotted as a bar plot (zoom in is shown on the upper left) within each circle (Cas9, red; eSpCas9, blue; Cas9 (ΔrecA), yellow). Genes with notable resistance to genome editing (the same threshold as in (B)) in each dataset (Cas9, eSpCas9 and Cas9 (ΔrecA)) are highlighted with gene names. Essential genes in rich medium are tagged with ‘(e)’. The heatmap (black to white) below the relevant bar plot of each circle indicates the standard deviation of the within-gene sgRNA activity for each highlighted gene. The color bar is shown on the right. A high-resolution version of this genome-wide map is accessible (https://figshare.com/s/127cecee6f9ea4e814e2) for downloading.