Expression of the cryIB crystal protein gene of Bacillus thuringiensis

The cryIB gene of Bacillus thuringiensis subsp. thuringiensis HD-2 codes for a Mr 139492 protein that is lethal to certain lepidopteran larvae. We used primer extension to map transcriptional initiation sites and found that cryIB was transcribed from two sites that are activated at different times during sporulation. The presumed promoter regions for the two start sites are very similar to the two promoters preceding the cryIA (a) gene, and the in vivo transcriptional start sites were found to be identical. Variable amounts of the full-length cryIB protein were detected by immunoblotting of extracts of recombinant cells of Escherichia coli; larger amounts were found when the TTG translational start codon was changed to ATG and when an htpR- strain of E. coli was used as the recipient for transformation. When expressed in E. coli, the cryIB protein was found to be toxic to the larvae of Artogeia rapae (LC50 of 58 ng/cm2) and exhibited little toxicity to the larvae of Manduca sexta (LC50 greater than 5000 ng/cm2).