FIGURE 6.

Identification of a promoter within the sbnC coding region. A, DNA fragments, spanning genes sbnA–E, were cloned into the pGylux vector, and luciferase activity was measured. Activity was expressed at cps/A₆₀₀. Statistical analyses were performed for each respective fragment, comparing the luciferase activity of either the sbnI strain without iron (sbnI − Fe) to WT without iron (WT − Fe) or WT with iron (WT + Fe) to WT without iron (WT − Fe). Statistics were performed using Student's unpaired t test; *, p < 0.01, ns = not significant. B, identification of initiation of transcription. Using the 5′RACE method, A81175 (based on S. aureus Newman genome sequence) was identified as the +1 site and is located 625-bp downstream of the sbnC start codon. Two PCR products of ∼760 bp (AUP/GSP1) and 117 bp (abridged universal anchor primer/GSP2) were generated, and the 117-bp product was cloned into pGEM7 and sequenced. The locations of the primer GSP2 and the −10 and −35 sites are indicated.