Molecular Cloning and Expression of the Chitinase Gene from Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus in E.coli

WU Dong; W ANG Hua—lin; DENG Fei; CHEN Xin—wen; PENG Hui—yin。HU Zhi—hong

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August 2002

WU Dong, W ANG Hua—lin, DENG Fei, CHEN Xin—wen, PENG Hui—yin。HU Zhi—hong and Molecular Cloning and Expression of the Chitinase Gene from Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus in E.coli[J]. Virologica Sinica, 2002, 17(4): 331-335.

Citation: WU Dong, W ANG Hua—lin, DENG Fei, CHEN Xin—wen, PENG Hui—yin。HU Zhi—hong, . Molecular Cloning and Expression of the Chitinase Gene from Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus in E.coli .VIROLOGICA SINICA, 2002, 17(4) : 331-335.

棉铃虫单核衣壳核多角体病毒几丁质酶基因的克隆与表达

1.
中国科学院武汉病毒研究所无脊椎动物病毒学联合实验室，湖北武汉430071

摘要

根据棉铃虫单核衣壳核多角体病毒(HaSNPV)几丁质酶基因的序列，设计引物，引入适当的酶切位点．利用 PCR扩增出不含N末端信号肽以及C末端内质网定位肽的几丁质酶基因片段。将该基因片断克隆至原核表达载体pProEXHTb，经IPTG诱导，在大肠杆菌DH5a中获得了高效表达，表达产物的大小为60kD，含量占菌体总蛋白量的4O％。利用来源于AcMNPV几丁质酶的抗体对表达蛋白进行检测，获得特异性的显色信号，证实所获原核表达产物与杆状病毒的几丁质酶具有同源性。
- 棉铃虫单核衣壳核多角体病毒
- , 几丁质酶
- , 克隆
- , 表达

Molecular Cloning and Expression of the Chitinase Gene from Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus in E.coli

1.
Joint—lab of Invertebrate Virology，Wuhan Institute of Virology，Chinese Academy of Sciences，Wuhan 430071．China

Abstract

The PCR product of the HaSNPV chitinase gene，which was without the N—terminal signal peptide and the C-terminal endoplasmic reticulum location signal，was cloned into a prokaryotic expres— sion vector pProEXHTb．After the induction of IPTG，the chitinase gene was successfully expressed in Eschechia coli DH5a．The expression product shown a molecular weight of 60 kDa，and it covered about 40％ of the total protein in E ．coli．W estern blot analysis using an antibody derived from AcM — NPV ehitinase confirm ed the expression product was a homologue of baculoviral chitinase．
- Helicoverpa armigera single—nucleocapsid nucleopolyhedrovirus
- , Chitinase
- , Cloning
- , Expression

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Molecular Cloning and Expression of the Chitinase Gene from Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus in E.coli

1. Joint—lab of Invertebrate Virology，Wuhan Institute of Virology，Chinese Academy of Sciences，Wuhan 430071．China

Abstract: The PCR product of the HaSNPV chitinase gene，which was without the N—terminal signal peptide and the C-terminal endoplasmic reticulum location signal，was cloned into a prokaryotic expres— sion vector pProEXHTb．After the induction of IPTG，the chitinase gene was successfully expressed in Eschechia coli DH5a．The expression product shown a molecular weight of 60 kDa，and it covered about 40％ of the total protein in E ．coli．W estern blot analysis using an antibody derived from AcM — NPV ehitinase confirm ed the expression product was a homologue of baculoviral chitinase．