Analysis of the effect of a pnp deletion and the rneΔ14 and rne-131 alleles on positive regulation of RpoS expression by DsrA and RprA. Western blot analysis of RpoS from exponential phase cultures of isogenic pnp+ and Δpnp strains either wild-type for rne (DJ624 and NRD579) or carrying rneΔ14 (NRD475 and NRD589) or rne-131 (NRD476 and NRD578) harboring either a vector or a plasmid that expresses DsrA (A) or RprA (B) from a lac promoter. Overnight cultures were diluted 200-fold in fresh LB medium with ampicillin, growth at 37°C to an OD600 of 0.3–0.4, the sRNA was induced by addition of IPTG for 20 min. Samples were then taken, the protein TCA-precipitated, and an equal amount of protein based on the cell density of each culture processed as described in Materials and Methods and developed using an anti-RpoS polyclonal antibody. The band intensity from each lane was quantified using the Multi Gauge software. The band intensity for the wild-type strain was set to 100%, and other samples were normalized to the wild-type strain. These results represent the mean of two experiments.
