beta-Glucuronidase from Escherichia coli as a gene-fusion marker.

Abstract

We have developed a gene-fusion system based on the Escherichia coli .beta.-glucuronidase gene (uidA). The uidA gene has been cloned from E. coli K-12 and its entire nucleotide sequence has been determined. .beta.-Glucuronidase has been purified to homogeneity and characterized. The enzyme has a subunit molecular weight of 68,200, is very stable, and is easily and sensitively assayed using commercially available substrates. We have constructed gene fusions of the E. coli lacZ promoter and coding region with the coding region of the uidA gene that show .beta.-glucuronidase activity under lac control. Plasmid vectors have been constructed to facilitate the transfer of the .beta.-glucuronidase coding region to heterologous control regions, using many different restriction endonuclease cleavage sites. There are several biological systems in which uidA-encoded .beta.-glucuronidase may be an attractive alternative or complement to previously described gene-fusion markers such as .beta.-galactosidase or chloramphenicol acetyltransferase.