FIG. 7.

Primer extension analysis of the promoter fixRp₁- and fixRp₂-dependent fixR′-′lacZ transcripts in wild type and ΔUAS and regR mutants containing a chromosomally integrated fixR′-′lacZ fusion. Total RNA was purified from the indicated B. japonicum strains grown aerobically (+O₂) in PSY medium or anaerobically (−O₂) in YEM medium plus KNO₃. Hybridization to RNA with the ³²P-labeled oligonucleotides lac4B and PBj16S and reverse transcription of the fixR′-′lacZ mRNA and the 16S rRNA primary transcript, respectively, were performed as described in Materials and Methods. The products were separated on a 6% polyacrylamide gel next to a sequence ladder of plasmid pRJ7211 made with oligonucleotide lac4B. Transcripts T1, T2, and Bj16S (control) are marked. The origin of the unmarked reverse transcription products present in all lanes is not known; they had not been observed in similar, previous studies in which a shorter lacZ-specific oligonucleotide (lac4 [5]) was used.