Expression profiles of trans -encoded sRNAs in response to antibiotics and serum. (A) Northern blot analysis was performed on 6 μg total RNA isolated from Bt CDC272 grown at 37°C to mid-exponential (OD600 ~ 0.5, left panel) or early stationary phase (OD600 ~ 0.8, right panel) in LB or LB containing 300 μg/ml gentamycin (LB + Gent), 200 μg/ml kanamycin (LB + Kan), or 20% bovine serum (LB + Serum). At OD600 ~ 0.5 and ~0.8, the fold change in BTH_s1 and s39 transcript levels (displayed under the RNA bands) was determined as the ratio of sRNA normalized to 5S rRNA for each experimental condition, compared to growth in LB. To evaluate the effect of signal normalization between independent probe annealing experiments, the left panel includes RNA samples from bacteria grown in LB to OD600 ~ 0.8 (left panel, first lane) for comparison to transcript levels at OD600 ~ 0.5. One representative of four Northern blots performed on RNA isolated from independent experiments is shown. (B) Two-step RT-PCR analysis was performed on total RNA isolated from Bt CDC272 grown at 37°C to OD600 ~ 0.5 (left plot) and OD600 ~ 0.8 (right plot). The relative RNA levels are presented as fold change in BTH_s1, s39, and s36 expression levels between LB + treatment compared relative to LB alone. For each sample, the sRNA levels were normalized to 5S rRNA. The amount of antibiotic and serum used was as described in (A). The data represents the average and standard deviation from three independent experiments performed in duplicate. The “*” denotes statistical significance (p < 0.05) between sRNA expression in B. thailandensis grown in LB + treatment compared to LB alone.