Genotyping and Molecular Characterization of Local Bacillus thuringiensis Isolates of DOR by using PCR Based Methods

Not AvailableEight Bacillus thuringiensis (Bt) strains were isolated from the soil samples collected from different fields in Andhra Pradesh and Karnataka, India. The isolates were characterized on the basis of primary bioassay against 2nd instars larvae of Helicoverpa armigera and Spodoptera litura and also screened with REP-PCR (Repetitive Extragenic Palindromic sequence) to distinguish closely related relationship among the strains. Based on the bioassay of eight isolates, Bt-52 and Bt-127 isolates were showed high toxicity against 2nd instars of H. armigera and S. litura. Rep-PCR profiles with these isolates showed Bt-52 and Bt-127 isolates formed separate clusters in phylogenetic tree. Further PCR amplification was carried out with cry gene specific primers used for 8 Bt isolates revealed the presence of amplified fragments characteristic of ten different cry genes. Results showed the expected sizes of PCR products of cry1Aa, cry1Ac, cry1Ad, cry1Ae, cry2Aa, cry2Ab, cry2Ac, cry1C, cry1D and cry1E genes ranging from 1000-1286, 844, 500, 443 and 300 bp, respectively. Out of the eleven cry genes investigated, only cry1Ab gene was not found any of the 8 Bt isolates. One Bt isolate Bt-112 showed only cry2Aa gene with PCR product size of 500bp. While, six of the 8 isolates were harboring a combination of different cry genes, which are Lepidoptera active cry1 genes and Lepidoptera-Diptera active Lepidoptera active cry2 genes. Consequently, combined strains are good candidates in the search for bio-control agents with a wider spectrum of action against many different insects. On the other hand, cry1 genes were the most frequently found among the 8 local Bt isolates and the most common profile of cry2 genes contained cry2Aa and the second abundant genes are cry1Ad, cry2Ab, cry2Ac and cry1C.Not Availabl