Cloning, Expression And Identification The Mosquitocidal Of The Cyt2B Gene In Bacillus Thuringiensis LLP29

Cyt toxin is produced by Bacillus thuringiensis.subsp. israelensis (Bti) during sporulation. Based on the cyt2B sequence reported in the NCBI, this study constructed two degenerated primers. Using these primers and the total DNA of the Bt LLP29 as a template, a 1,082 bp DNA fragment was amplified by polymerase chain reaction (PCR) and cloned into E. coil JM109. After sequencing, the cyt2B protein was produced in E. coil BL21 using pGEX-KG as expression vector.The nucleotide sequence showed two open reading frames (ORF) in the cyt2B fragment. One was 789 bp long, encoding a protein of 262 amino acide. It is very similar in size to the reported Cyt2B protein. The homology search of the cloning gene displayed that it had 98% similar to 5 reported genes which were reported as cyt2Ba7 or cyt2Ba8. SDS-PAGE analysis indicated that the Cyt2B protein had a molecular mass of about 30kD, similar to the reported Cyt2Ba protein which was previously been shown toxic to mosquitos. Toxicity activities of the recombinant strains were tested on Aedes albopictus and Culex pipiens fatigans. Compared with the wild Bt LLP29 strain, the recombinant strain didn't show any toxicing against them.