Figure 6. Screening mismatched sgRNA libraries in combination with chemical and genetic perturbations reveals modulators of essential gene requirements.

(A) RT-qPCR measurements of repression by 2 fully complementary and 2 mismatched sgRNAs targeting 4 essential genes and rfp in E. coli shows that both mismatched and fully complementary sgRNAs are able to repress transcription. Mismatched sgRNAs (labeled *.mm) repress transcription less efficiently than their fully complementary counterparts. asd and glmS are less efficiently repressed by all 4 sgRNAs (both fully complementary and mismatched), suggesting that transcriptional or post-transcriptional feedback is important in regulating their level. (B) Schematic of peptidoglycan recycling and synthesis pathway in E. coli. (C-D) Volcano plots comparing the median change in relative fitness for all sgRNAs targeting a gene to the statistical significance of those changes as quantified by a Wilcox test in the ΔampG (C) and Δmpl (D) genetic backgrounds. The dashed line represents a Bonferroni corrected p-value < 0.01