Cloning and expression product of vip3A gene from Bacillus thuringiensis and analysis of inseceicidal activity.
- Author:
Jian-Wu CHEN
1
;
Li-Xia TANG
;
Mu-Jin TANG
;
Yong-Xia SHI
;
Yi PANG
Author Information
1. State Key Laboratory for Biocontrol, Zhongshan University, Guangzhou 510275, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Bacillus thuringiensis;
genetics;
Bacterial Proteins;
genetics;
isolation & purification;
pharmacology;
Base Sequence;
Cloning, Molecular;
Escherichia coli;
genetics;
Insecticides;
pharmacology;
Molecular Sequence Data;
Pest Control, Biological;
Recombinant Proteins;
biosynthesis;
isolation & purification;
pharmacology;
Spodoptera
- From:
Chinese Journal of Biotechnology
2002;18(6):687-692
- CountryChina
- Language:Chinese
-
Abstract:
The vip3 A gene in a size of 2.3 kb amplified from wild-type Bacillus thuringiensis strain S184 by PCR was cloned into pGEM-T Easy vector and its sequence was analysized by DNASTAR. The plasmid pOTP was constructed by inserting vip3A-S184 gene into the expression vector pQE30 and then was transformed into E. coli M15. E. coli M15 cells harbouring the plasmid pOTP were induced with 1 mmol/L IPTG to express 89 kD protein which was confirmed to be Vip3A-S184 by Western blot. Experiments showed that about 19% of Vip3A-S184 proteins were soluble, and others were insoluble proteins and formed inclusion bodies observed by transmission electron microscopy(TEM). The target protein was purified under the native condition and the polyclonal antibody was prepared by immunizing rabbits. The polyclonal antibody was used to detect Vip3A proteins expressed in Bacillus thuringiensis. Bioassay showed that Vip3A-S184 showed a high toxicity against 3 tested insect larvae including Spodoptera exigua, Spodoptera litura and Helicoverpa armigera.
