Effect of promoter substitution on amino acid and glucose repression. Starch diffusion plate assay and western blot analysis of AmyA levels culture supernatant grown in a medium with Tryptone. Panel A, no treatment. Panel B, Iodine treatment. Numbered spots were: 1 (wild type, PBL2025); 2 (malAp-amyA, PBL2058); 3 (amyA::lacS, PBL2004). All spots contained 107 cells and plates were incubated at 80 °C for 6 days. Panel C, three-fold serial dilutions of wild type (PBL2025) culture supernatant and three-fold serial dilutions of the malAp::amyA promoter fusion strain (PBL2058) culture supernatants. In Panel C, purified AmyA standard (STD, 40 ng) is indicated. Lane 1 in both panels contains protein dervived from a culture volume proportional to 109 cells. Panel D, western blot analysis of culture supernatants from PBL2025 (wild type) and PBL2058 (malAp::amyA) strains grown in a medium containing either glucose (Glc) or starch (St) as sole carbon and energy sources respectively. Samples of culture supernatants were equivalent to a volume proportional to 5 × 109 cells. Panel E, western blot analysis of three-fold serial dilutions of supernatants from PBL2025 (wild type) grown in glucose (lanes 1–3) or starch (lanes 4–6) as sole carbon and energy sources. Purified AmyA (STD, 40 ng) was used as a standard. Lanes 1 and 4 contain protein dervived from a culture volume proportional to 6 × 108 cells.