Cloning of a New Truncated cry1Ac Gene from an Indian Isolate of Bacillus thuringiensis

Transgenic Bt crops producing insecticidal crystal proteins from Bacillus thuringiensis (Bt), so-called Cry toxins, have proved useful in controlling insect pests. Among the cry toxins, Cry1A toxins are important because of high toxicity to lepidopteran pests and their widespread distribution among Bt strains. In Cry1A proteins, toxin fragment is comprised of about 620 amino acids of N-terminal region and C-terminal half is not required for toxicity. Four indigenous isolates of Bt viz., T15, T16, T20 and T31 were screened by PCR-RFLP for 3'-truncated cry1A gene(s) corresponding to toxin fragment. RFLP analysis of cry1A amplicons obtained from the four isolates of Bt showed presence of cry1Ac-type gene alone in three isolates. One of the cry1Ac-postive isolates, T15 which showed 100 percent mortality in Helicoverpa armigera, was selected for cloning of DNA fragment of about 2.1 kb containing 3'-truncated cry1Ac gene. Nucleotide sequence data generated for 3'-truncated cry1Ac gene of T15 showed 98 to 99 percent homology with 1958 bp of already reported sequences of all cry1Ac genes (cry1Ac1 to cry1Ac24). Deduced amino acid sequence of cry1Ac of Bt strain, T15 showed one to four percent variation in comparison to all reported Cry1Ac holotypes (Cry1Ac1 to Cry1Ac24) by differing at 5 to 19 positions. This suggests that the cry1Ac toxin of Bt isolate, T15 is a new kind of its group.