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| Gene Synthesis of Bacillus thuringiensis Cry3Aa and High-level Expression in Burkholderia pyrrocinia JK-SH007 as a Biopesticide |
| LI Yang1, WU Chou-fei2, WU Xiao-qin1, YE Jian-ren1, ZHANG Li-qin1 |
1. College of Forest, Nanjing Forestry University, Nanjing 210037, China;
2. College of Life Sciences, Huzhou Teachers College, Huzhou 313000, China |
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Abstract To develop an anti-Pyrrhalta larva biopesticide. The T7 RNA pol for T7 DNA-directed RNA polymerase was cloned from BL21(DE3) into the Burkholderia pyrrocinia expression vector pHKT2. The insecticidal cry3Aa gene from Bacillus thuringiensis was optimized, synthesized, and cloned to the modified T7 expression system. The expression of the cry3Aa gene in wild-type endophyte B. pyrrocinia JK-SH007 was induced with temperature shifting, resulting in high-level production of Cry3Aa protein with a molecular weight of about 70 kDa. Toxic mortality bioassay was conducted on 15 P. aenescens(Fairmaire) larvae in the 2 instar in the laboratory. The crude engineered strain was highly toxic[LC50=0.63(0.48~0.82) g/L] to Pyrrhalta aenescens(Fairmaire) larvae grown in culture dishes on elm leaves. In conclusion, a novel T7 expression vector for the effective expression of a cry3Aa gene for a protein toxic to beetle larvae was devised, and its impactful delivery can make it a possible candidate for a biopesticide.
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Received: 15 March 2017
Published: 25 June 2017
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