In vitro transcription of the cloned chromosomal crystal gene from Bacillus thuringiensis

We have determined the conditions required for in vitro transcription of the cloned chromosomal crystal gene from Bacillus thuringiensis using either the homologous vegetative RNA polymerase or a sporulation specific form of this enzyme. The gene is actively transcribed by the latter enzyme (form II) but not by the vegetative one. Evidence for a specific recognition between the form II enzyme and the promotor site of the crystal gene was obtained by binding experiments. They showed that the binding is increased by the presence of some additional factors, which change the specificity of the vegetative core-enzyme. The sequence of the promoter has been determined and the start-point of the transcription deduced. Two hexanucleotide sequences, TACAAT and CCTACG, centered at - 10 and - 35 bp are present, but are somewhat different from the consensus sequences previously described in other bacilli.