The effect of the clpP, rpoS and csrA genes on the level of RpoS and expression of csrA . Cells were grown to late log phase (OD600 of 0.65) in LB at 37°C or cold-shock at 15°C. A) The level of RpoS determined by Western blot in the wild type, clpP mutant and six cold resistant clpP suppressor mutants isolated from three independent experiments. Suppressor 1.1 and 1.2 was from the initial isolation of 12 random isolated. Suppressor 2.1 and 2.2 was from the quantification of suppressor frequency. Suppressor 3.1 and 3.2 was isolated at day 14 from other biological replicate of growth at 10°C. B) The level of RpoS determined by Western blot in the wild-type C5 and isogenic mutants before and after 3 hours of cold shock. C) The expression of csrA in the wild type and clpP, rpoS, csrA (sup) and clpP/rpoS mutants. RNA was extracted, dot blotted onto a hybridization filter and hybridized with labelled csrA probe. All figures are images of one representative gel where irrelevant samples have been edited away.