Two-vector assay as a tool for examining Spo0A gene transcription regulation

We have modified an assay using two compatible vectors that coexist in Escherichia coli cells and applied it in the investigation of the transcriptional activity of Spo0A, a key regulator of sporulation in Bacillus subtilis. We have chosen the promoters of the Spo0A dependent genes, spoIIE and spoIIA, involved in sporulation, in order to study the transcription activity solely of the DNA binding domain of Spo0A. We have prepared the two-vector system so that one vector contained the cloned C-Spo0A under the control of an inducible promoter, and the second vector (the promoter probe vector), was composed of the Spo0A dependent spoIIE and spoIIA promoters. Using this two-vector system in E. coli, we proved that C-Spo0A is able to interact with the E. coli transcription apparatus, recognizes both promoters and activates transcription from these promoters.