FIG. 2.

Activity of the cloned Anabaena RNAP on the promoter of the psbA gene. (A) Results of DNase I footprinting. The DNA fragment of the promoter region of the psbA gene was incubated without (lanes 1 and 3) or with (lane 2) purified RNAP holoenzyme (11 nM plus 40 nM SigA) before DNase I treatment (noncoding strand represented). The positions of the −10 and −35 determinants as well as the footprinted region are indicated by vertical lines. (B) Results of potassium permanganate footprinting. The DNA fragment encompassing the promoter of the psbA gene was incubated with purified RNAP holoenzyme (60 nM plus 100 nM SigA) before potassium permanganate treatment (lane 1, the coding strand subjected to primer extension). The −10 promoter element is indicated in the sequence ladder. (C) Results of the in vitro runoff transcription assay. The psbA DNA fragment was incubated without (lane 1) or with (lane 2) RNAP (15 nM plus 48 nM SigA) before the addition of nucleoside triphosphates (see Materials and Methods for details). The arrowhead indicates the full transcript corresponding to the DNA fragment used. Size markers (in nucleotides) are also indicated.