FIG. 3.

Identification of the minimal region of the davD promoter that allowed regulated expression. Intergenic regions of different length between davD and orf3 were amplified by PCR with the same primer to meet the 3′ end (provided with a Kpn site) and different primers at the 5′ end (provided with an EcoRI site). The PCR products, once digested with these restriction enzymes, yielded fragments of 416, 280, 137, 81, and 26 bp upstream from the +1 transcription start point. P. putida KT2440 cells bearing the indicated fusion were grown on M9 minimal medium with benzoate alone or with 5 mM lysine or 5 mM δ-aminovaleric acid. The β-galactosidase activity was determined as described in Materials and Methods, and values are the averages of three independent assays done in duplicate.