FIG 3.
Stringent rpoB mutations rescue the inviability of a Plac-rne strain grown without inducer (IPTG). (A) Wild-type and rpoB mutant derivatives of Plac-rne strains were streaked on a pair of LB plates with the indicated micromolar IPTG concentrations and incubated at 37°C until the growth on plate without IPTG was clear (for 2 days). The strains used, from left to right, were GJ6974, GJ17402, GJ17403, GJ17404, and GJ17405. (B) Cultures of strains GJ17403, GJ17405, and GJ6974 were grown in LB with IPTG (100 μM), washed, and plated at suitable dilutions on LB plates with the indicated IPTG concentrations, and RNase E levels in the cells of colonies harvested from the plates after 48 h were determined by Western blotting. In the upper panel, the RNase E band is marked by the arrow; the lower panel shows a representative section of the same blot stained with amido black, which serves as a loading and transfer control. The band seen immediately below that of RNase E in this experiment was inferred to be a nonspecific band, since it is also observed in the lane for RNase E-ΔCTH (see also Fig. S4 in the supplemental material).