ABSTRACT
Discovery of novel cry genes of Bacillus thuringiensis (Bt) with higher toxicity is important for the development of transgenic Bt crops resistant to target pests. Two new indigenous isolates of Bt were characterized for their colony type, crystal inclusion and toxicity with Helicoverpa armigera Hubner and Spodoptera litura Linn. Screening of cry2A genes from the two new isolates of Bt, T30 and T48, by PCR with cry2A family primers showed the presence of cry2A genes only in the isolate T30. Further screening of these isolates of Bt with three different cry2A gene specific primers showed the presence of cry2Aa, cry2Ab and cry2Ac genes in both of the new Bt isolates. The cry2A open reading frame (ORF) of Bt T48 (~1.9 kb) was amplified and cloned in a T/A vector (~2.8 kb). All the E. coli transformants showed only the cry2Aa gene. Comparison of the nucleotide sequence data generated from the cry2Aa (~1.9 kb) gene showed a 99 percent homology and one amino acid variation when compared with the holotype sequence of Cry2Aa1.
ABSTRACT
The Western Ghats of India is the one of the world's 10 "Hottest biodiversity hotspots" that runs along the western part of India through four states including Tamil Nadu. The only biodiversity reserve in the Western Ghats is the Nilgiri biosphere located in the Tamil Nadu state. In the present study, 525 soil samples were collected from all the 14 different divisions of the Western Ghats in Tamil Nadu state, India. A total of 316 new isolates of Bacillus thuringiensis (Bt) that produce parasporal crystalline inclusions were isolated from 525 soil samples. Seven different types of crystalline inclusions were observed in the 316 new isolates of Bt. Cuboidal inclusion was predominantly present in 26.9% of the Bt isolates when compared to other shapes. Further characterization of 70 of the 316 Bt isolates for crystal protein profile through SDS-PAGE revealed six different types of crystal protein profile viz., 135 and 65, 135, 95, 65, 43, and 30 kDa crystal proteins. Variation in the mass of crystal protein(s) purified from the isolates of Bt revealed molecular diversity of this bacterium prevalent in the Western Ghats of Tamil Nadu, India.
