Fig. 1: A SoxS double mutant maintains CRISPRa activity and does not activate endogenous SoxS targets. | Nature Communications

Fig. 1: A SoxS double mutant maintains CRISPRa activity and does not activate endogenous SoxS targets.

From: Effective CRISPRa-mediated control of gene expression in bacteria must overcome strict target site requirements

Fig. 1

a Reporter system for measuring the CRISPRa activity and endogenous SoxS-dependent gene expression of wild-type or mutant SoxS constructs. CRISPRa activity was determined in a strain harboring a genomically integrated sfGFP reporter (CD06, Supplementary Table 1). The endogenous SoxS-dependent gene expression was determined by monitoring lacZ expression from reporter plasmids where lacZ was driven by SoxS-regulated promoters zwfp and fumCp44. GFP fluorescence was measured by flow cytometry and lacZ activity was measured using a β-galactosidase assay. b SoxS(R93A/S101A) maintains CRISPRa activity and does not activate expression from the endogenous expression from the zwfp and fumCp reporters. Fluorescence and lacZ activity values were baseline-subtracted using a strain that does not express a scRNA. Both GFP levels and lacZ activities were normalized to the values observed in the strain with wild-type SoxS. Values represent the average ± standard deviation calculated from n = 3 biologically independent samples. Source data of b are provided as a Source Data file.

Back to article page