CLONING AND CHARACTERIZATION OF CRY1 AND CRY2 GENESFROM INDIGENOUS BACILLUS THURINGIENSIS ISOLATESSantosh Kumar D., S. Ramgopal Rao, and Dwarkesh S. PariharAbstract Insecticidal crystal proteins produced by Bacillus thuringiensis (Bt) are toxic to specific insect pests. They areused as alternatives to chemicals for controlling insect pests using transgenic plants and biopesticides.Three strains of Bacillus thuringiensis expressing cry1 and cry2 genes were identified in soil samples collectedfrom different locations in India. The full-length cry1, cry1I, and cry2 type genes (3.5, 2.1, and 1.9 kb,respectively) were cloned and sequenced. The full-length clones of the three genes were sub-cloned in theE. coli expression vector, and expressed proteins were analysed. The expression of cry1, cry1I, and cry2resulted in 132, 81, and 70 kDa proteins, respectively. The expressed proteins were tested against secondinstarlarvae of Helicoverpa armigera. The expressed proteins of cry1andcry2- type from BRI-86 exhibited thehighest toxicity to H. armigera larvae with an LC50 of 2.34 and 0.2 μg/ml diet, respectively. The LC50exhibited by the test proteins was two and four-fold lower than the reference Btk HD-1 proteins. Thus, thespore preparation or the genes can be used individually or in combination to control insect pests. |