Figure 3.

In vitro transcription from esp1396I promoters. (A) Escherichia coli RNAP σ⁷⁰ holoenzyme (100 nM) was combined with DNA fragments containing 8 nM Pesp1396ICR (A–C) or Pesp1396IM (D and E) in the presence, where indicated, of C.Esp1396I, whose concentrations ranged 0 to 500 nM. In (D), a component indicated by red color was added to DNA first, following an incubation long enough to allow complex formation and subsequent addition of component indicated by black color. In (A and D), complexes were probed with KMnO₄. On the left of these panels, the sequence of the relevant portion of each promoter is mapped on the A+G sequencing marker lane. In (B and E), a single-round transcription reaction was performed. In (C), abortive initiation reaction was performed using RNAP holoenzymes reconstituted with indicated σ factors. Reaction products were separated by denaturing gel electrophoreses and revealed using a Phosphorimager.