Cloning and endonuclease restriction analysis of uidA and uidR genes in Escherichia coli K-12: determination of transcription direction for the uidA gene

Abstract

The two genes of the Escherichia coli K-12 uid region (the structural gene uidA and the regulatory gene uidR) were isolated on a ColE1-uid hybrid plasmid from the bank of Clarke and Carbon. We made a restriction map of this region and correlated it with the genetic map by subcloning the uid restriction fragments into plasmids pBR322, pBR325, and pACYC177. In these plasmids, amplification of the products of the uidA and uidR genes occurred. The enzyme coded for by uidA was identified by polyacrylamide gel electrophoresis of crude extracts from strains containing the uidA plasmid. A 1-megadalton EcoRI-BamHI segment contained the uidAo operator, and the direction of transcription of the uidA gene was determined. The restriction analysis also suggested that the order of the loci in this region is manA, uidA, uidAo, uidR.