Figure 1.
Toxicity of [C2mim]Cl to E. coli DH1 upon addition of [C2mim]Cl during exponential growth.
Red: 0 mM [C2mim]Cl, purple: 50 mM [C2mim]Cl, blue: 100 mM [C2mim]Cl, green: 150 mM [C2mim]Cl, yellow: 200 mM [C2mim]Cl, orange: 400 mM [C2mim]Cl. Error bars represent standard errors.
Table 1.
Comparison of microarray and qPCR results for selected genes after stress with 150[C2mim]Cl.
Figure 2.
qPCR analysis of inducing PydfA’, PydfO’ and PmarR’ in E. coli DH1 in 150 mM [C2mim]Cl, 100 mM [C2mim][CH3COO], 150 mM NaCl and 100 mM Na[CH3COO].
Cultures were grown until mid-exponential phase, when the compounds were added and RNA samples were collected after 30 min. The fold change on the chart represents the change in expression compared to an untreated control. hcaT was used as an endogenous reference to normalize the data. Error bars represent standard errors. Blue: 150 mM [C2mim]Cl, red: 100 mM [C2mim][CH3COO], green: 150 mM NaCl, purple: 100 mM Na[CH3COO].
Figure 3.
Transcript levels of ydfO, ydfA and marR during exponential and stationary growth after exposure to [C2mim]Cl.
[C2mim]Cl was added from the beginning of growth at the indicated concentrations, and cultures were harvested during exponential growth (OD600 = 0.5) and stationary growth. The fold change represents the change in expression compared to an untreated control. hcaT was used as an endogenous reference to normalize the data. Error bars represent standard errors. Blue: 50 mM [C2mim]Cl, red: 100 mM [C2mim]Cl, green: 150 mM [C2mim]Cl, purple: 200 mM [C2mim]Cl.
Figure 4.
Growth of different eilA expression strains at increasing [C2mim]Cl concentrations.
A–E: growth assays of E. coli DH10B carrying different promoter-eilA constructs. Due to day-to-day variability in the final cell density reached in the microtiter-plate experiments, growth curves were normalized to a start OD of 0 and a maximum OD of 0.5. A) pPydfO’-eilA, B) pPydfA’-eilA, C) pPmarR’-eilA, D) pPlacUV5-eilA, E) pP-eilA. Error bars represent standard errors. Blue: 0 mM [C2mim]Cl, red: 100 mM [C2mim]Cl, green: 200 mM [C2mim]Cl, purple: 400 mM [C2mim]Cl.
Figure 5.
Relative abundance of eilA expression strains after competitive growth in pools with increasing [C2mim]Cl concentrations.
Cultures were pooled and grown over 48-specific primers. 10 µM IPTG was added to induce the pPlacUV5-eilA construct. qPCR results were normalized using cat (the gene conferring chloramphenicol resistance on the expression plasmid) as endogenous control. Error bars represent standard errors. Dark blue: pPlacUV5-rfp, red: pP-eilA, green: pPydfO’-eilA, purple: pPydfA’-eilA, light blue: pPmarR’-eilA, orange: pPlacUV5-eilA.
Figure 6.
Quantification of EilA protein levels in different growth phases at increasing concentrations of [C2mim]Cl.
Strains contain eilA expression constructs are as indicated in the graph. EilA protein levels were quantified during exponential (A) and stationary (B) growth, at the indicated [C2mim]Cl concentrations. EilA protein levels were normalized against Cat protein levels as an endogenous control. No growth was observed for pP-eilA and pPydfO’-eilA in the presence of 400 mM [C2mim]Cl. Error bars represent standard errors. Blue: 0 mM [C2mim]Cl, red: 50 mM [C2mim]Cl, green: 150 mM [C2mim]Cl, purple: 400 mM [C2mim]Cl.
Figure 7.
Modeling results comparing controllers at different [C2mim]Cl concentrations.
A) pPydfO’-eilA, B) pPydfA’-eilA, C) pPmarR’-eilA, D) pPlacUV5-eilA, and E) pP-eilA. Blue: 0 mM [C2mim]Cl, red: 100 mM [C2mim]Cl, green: 200 mM [C2mim]Cl, purple: 400 mM [C2mim]Cl.
Table 2.
Plasmids used in this study.
Table 3.
Numerical constants used in simulations.