Figure 4

Analysis of pgaABCD regulation by PNPase. A. Northern blot analysis of pgaABCD operon transcription. 15 μg of total RNA extracted from E. coli C-1a ( pnp⁺) and E. coli C-5691 (Δpnp-751) cultures grown up to OD₆₀₀ = 0.8 in M9Glu/sup at 37°C were hybridized with the radiolabelled PGA riboprobe (specific for pgaA). B. Identification of in cis determinants of pgaABCD regulation by PNPase. Map of pJAMA8 luciferase fusion derivatives and luciferase activity expressed by each plasmid. Details about plasmid construction and coordinates of the cloned regions are reported in Methods and in Table 1. Construct elements are reported on an arbitrary scale. For relative luciferase activity (R.A.) in E. coli C-5691 (Δpnp-751) vs. E. coli C-1a (pnp⁺) strains, average and standard deviation of at least two independent determinations are reported. Although the absolute values of luciferase activity could vary from experiment to experiment, the relative ratio of luciferase activity exhibited by strains carrying different fusions was reproducible. The results of a typical experiment of luciferase activity determination are reported on the right.