Plasmids with the uidA reporter gene for the detection of promoters and transcription signals

Abstract

We have constructed promoter-probe plasmids, pNB4 and pNB5, based on the promoterless gene for beta-glucuronidase (uidA) of Escherichia coli. Unique restriction sites for EcoRI, SacI, KpnI, SmaI, XmaI, XbaI, SalI, SphI and HindIII in pNB4 and for HindIII, PstI and BglII in pNB5 were included upstream of the uidA structural gene. The usefulness of these plasmids was demonstrated by cloning the promoter-operator region of the E. coli uxaB gene. We observed that expression of the uxaB-uidA operon fusion followed the transcription-regulating properties of the uxaB promoter. Another construct, pNB2, can be used to detect operator and terminator signals. Recipient cells transformed with such recombinant plasmids can be revealed by growth on medium containing a chromogenic beta-glucuronidase substrate.