Isolation of Cry and Vip genes from native Bacillus thuringiensis

Bacillus thuringiensis (Bt) is a Gram positive spore-forming soil bacterium that produces parasporal crystal proteins or d-endotoxin proteins (encoded by the cry genes) and cytolytic proteins (encoded by the cyt genes) during sporulation specifically toxic to many agriculturally important insect species. Another novel class of insecticidal proteins secreted during the vegetative phase includes vegetative insecticidal proteins (vip) and secreted insecticidal protein (sip). In the present study, presence of lepidopteran specific cry1, cry3 and vip3A type genes was investigated by polymerase chain reaction (PCR) using specially designed gene specific primers. The cry1 gene family was found to be present in ten out of twenty four native Bt isolates, while none of the isolates showed the presence of cry3 gene family and vip3 genes. A full-length 3.5 Kb cry1Ac type of gene was amplified from native Bt isolate SK-729 and sequenced. The sequence of the cloned gene was 99% in identical tothat of the previously reported cry1Ac2 gene except four nucleotides and two amino acids variation at the protein level. The sequence was deposited in the NCBI with accession. No DQ023296.