Engineered Cry1Ac-Cry9Aa hybrid Bacillus thuringiensis delta-endotoxin with improved insecticidal activity against...

Recombinant Bt construct was prepared by exchange of pore forming domain I with cry1Ac to cry9Aa gene by overlap extension PCR (OE-PCR) technique. Construction of cry1Ac-cry9Aa was accomplished by six base pair homology at 3' ends of PCR products of domain I of cry1Ac and domain II and III of cry9Aa. The recombinant toxin was also modified by deletion of N-terminal alpha helix-1 of recombinant toxin. Both Cry toxins were expressed in E. coli BL21(DE3) plysS and purified by His-tag purification. Upon insect bioassay analysis against devastating crop pest Helicoverpa armigera, toxicity of recombinant toxin was found around fivefold higher than native Cry1Ac while alpha helix-1 deleted N-terminal modified toxin did not resulted in significant increase in toxicity. The recombinant Cry toxins such as Cry1Ac-Cry9Aa and Cry1Ac-Cry9AaMod may be used for insect pest control.