Expression of orf1 from the Bacillus thuringiensis NRD-12 cry2Aa1 operon

The 5' untranslated region and the orf1 sequence from the cry2Aa1 operon from Bacillus thuringiensis subsp. kurstaki NRD-12 were sequenced and compared to that from strain HD-1. The start codon described in HD-1 does not yield in NRD-12 a protein of the expected size of 20 kDa, but a 10-amino acid peptide. A second, highly conserved start codon is located 25 bp downstream from the first one and corresponds to an open reading frame of the same size in all known orf1-related sequences. Expression of lacZ gene fusions created at the level of the first ATG, second ATG, and stop codon of the NRD-12 orf1 sequence showed that orf1 is translated from the second ATG. The expected protein is 19 kDa in size. The expression starts at t2, which is in agreement with the presence of a BtI promoter in the cry2Aa1 operon.