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. 1998 Aug;64(8):3092–3095. doi: 10.1128/aem.64.8.3092-3095.1998

FIG. 3.

FIG. 3

The rpoN null mutant is not affected in the transcription of nirS and norCB. Total RNA from MK516 and MK21 was prepared according to the method of Aiba et al. (1) from cells grown aerobically (lane 1 of each panel) or under denitrifying conditions (lane 2 of each panel). Induction of denitrification was as described in the text for the activity measurements. Samples (20 μg of RNA) were denatured by glyoxal-dimethyl sulfoxide treatment and separated on a 1.2% agarose gel (14). After transfer to a nylon membrane, the nirS and the norCB transcripts were detected by hybridization with digoxigenin-labeled probes (labeling kit from Boehringer Mannheim, following the instructions of the manufacturer). A 500-bp KpnI fragment of the nirS gene and a 2-kb PstI-BglII fragment of the norCB operon were used as probes. nirS exhibited mono- and polycistronic transcripts of 2 and 3.4 kb, respectively; the norCB transcript was 2 kb. Equal gel loading was verified by staining with acridine orange. The 16S and 23S rRNA species served as standards.