Locating essential Escherichia coli genes by using mini-Tn10 transposons: the pdxJ operon

Abstract

The mini-Tn10 transposon (delta 16 delta 17Tn10) confers tetracycline resistance. When inserted between a gene and its promoter, it blocks transcription and prevents expression of that gene. Tetracycline in the medium induces divergent transcription of the tetA and tetR genes within the transposon, and this transcription extends beyond the transposon in both directions into the bacterial genes. If the mini-Tn10 inserts between an essential bacterial gene and its promoter, the insertion mutation can cause conditional growth which is dependent on the presence of tetracycline. Two essential genes in adjacent operons of Escherichia coli have been detected by screening for tetracycline dependence among tetracycline-resistant insertion mutants. These essential genes are the era gene in the rnc operon and the dpj gene in the adjacent pdxJ operon. The pdxJ operon has not been described previously. It consists of two genes, pdxJ and dpj. Whereas the dpj gene is essential for E. coli growth in all media tested, pdxJ is not essential. The pdxJ gene encodes a protein required in the biosynthesis of pyridoxine (vitamin B6).