Kausar Malik

Genetic Improvement of a Local Isolate of Bacillus Thuringiensis Kausar Malik

A cry gene, obtained from a locally isolated Bt. strain, CAMB # 30382, highly effective against Tribolium castenium, was amplified through PCR. The gene was ligated to a cloning vector pGEM-T and was cloned into DH5?. Its characterization showed homology with coleopteran-specific cry3f gene from Bt. strain buibui, as judged by Southern hybridization, PCR, and Restriction analysis. The alignment of the sequenced gene with gene sequences present in gene data bank showed two silent mutations, at nucleotide 676 and 701. However, two other nucleotide changes at position 2887 and 2902 of holotype cry3f gene were resulting from amino acid change near 3? end were observed. These changes were causing change in the stretch of amino acids from valine, tyrosine, serine, glutamine & isoleucine in the holotype cry3f to glycine, phenylalanine, alanine, asparginine respectively in the clone from CAMB isolate 30382. This change may have drastic effect on the toxicity spectrum of the cry protein isolated from this local isolate in comparison with the cry protein from the Buibui. serovar japonesis. Partial sequencing has shown it to be a varient of cry3 gene, which may be a novel cry gene.