FIG. 1.

Expression of the G. stearothermophilus V ubiE gene in E. coli. Agarose gel (1.5%) electrophoresis of the RT-PCR products obtained using primers ORF600F and ORF600R, specific for the ubiE gene of G. stearothermophilus V, was performed (1). In each case, the template was 20 ng of total RNA extracted from E. coli AN70 (lane A), E. coli JM109 (lane B), E. coli AN70/600pr (lane C), or E. coli JM109/600pr (lane D). Lane E is a negative control to which no template was added. Std, molecular weight standard (HaeIII-digested Φ X174 replicative-form DNA). After incubating the RNA preparations at 25°C for 30 min with 1 unit of RQ1 DNase (Promega), the deoxyribonuclease was inactivated for 10 min at 70°C. The RT-PCR was then incubated for 1 h at 42°C and then for 30 cycles that consisted of 1 min at 95°C, 1 min at 55°C, and 1 min at 72°C, followed by a final extension of 10 min at 72°C.