Cloning and Expression of Nucleocapsid Protein Gene of Equine Arteritis Virus in E.coli

DU Jian; WANG Zhi-liang; ZHAO Yong-gang; SONG Hou-hui; JIN Ning-yi; ZHANG Nian-zu

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June 2005

DU Jian, WANG Zhi-liang, ZHAO Yong-gang, SONG Hou-hui, JIN Ning-yi and ZHANG Nian-zu. Cloning and Expression of Nucleocapsid Protein Gene of Equine Arteritis Virus in E.coli[J]. Virologica Sinica, 2005, 20(3): 323-325.

Citation: DU Jian, WANG Zhi-liang, ZHAO Yong-gang, SONG Hou-hui, JIN Ning-yi, ZHANG Nian-zu. Cloning and Expression of Nucleocapsid Protein Gene of Equine Arteritis Virus in E.coli .VIROLOGICA SINICA, 2005, 20(3) : 323-325.

马动脉炎病毒核蛋白基因的克隆与表达

杜建 ^1,3 ,
王志亮 ¹ ,
赵永刚 ¹ ,
宋厚辉 ⁴ ,
金宁一 ⁵ ,
张念祖 ^2*

1.
1.农业部动物检疫所国家外来动物疫病诊断中心山东青岛 2.云南昆明农业部热带亚热带动物病毒学重点实验室 3.昆明市畜牧兽医站 4.中国科学院微生物研究所 5.解放军军需大学病毒基因工程重点实验室北京吉林长春

摘要

马病毒性动脉炎是危害世界养马业的重要传染病之一,是由动脉炎病毒科动脉炎病毒属的马动脉炎病毒(Equinearteritisvirus,EAV)引起的一种以病马发热,步态僵直,躯干及眼周围水肿,并出现粘液脓性鼻炎、结膜炎,外生殖道水肿为特征的传染病,对妊马能引起流产,使易感怀孕母马的流产率?..
- 马动脉炎病毒
- , 核衣壳蛋白
- , 表达
- , 纯化

Cloning and Expression of Nucleocapsid Protein Gene of Equine Arteritis Virus in E.coli

Abstract

The gene of nucleocapsid protein of Equine arteritis virus was amplified from PMD-18-T plasmid with equine arteritis virus ORF7 sequence by PCR. The PCR product was sequenced as well as purified and digested with EcoR I and Xho I, then directly cloned into the prokaryotic vector pET32a. Consequently the recombinant plasmid was constructed, designateds pET32a-N. PET32a-N was transformed into the host cell BL21(DE3) and the expression procedure was optimized including cultivated temperature, optional induction concentration and time of IPTG. The result indicated that the nucleocapsid protein can be expressed efficiently with 0.8mmol/L IPTG and 4 hour induction. The resulting Trx-N recombinant fusion protein was identified to be consisted of 34 kDa protein by SDS-PAGE and western blotting analysis. It indicated that the recombinant fusion protein could be used as antigen of diagnostic assay for detecting antibodies.
- Equine arteritis virus
- , Neucleocapsid protein
- , 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

Cloning and Expression of Nucleocapsid Protein Gene of Equine Arteritis Virus in E.coli

Abstract: The gene of nucleocapsid protein of Equine arteritis virus was amplified from PMD-18-T plasmid with equine arteritis virus ORF7 sequence by PCR. The PCR product was sequenced as well as purified and digested with EcoR I and Xho I, then directly cloned into the prokaryotic vector pET32a. Consequently the recombinant plasmid was constructed, designateds pET32a-N. PET32a-N was transformed into the host cell BL21(DE3) and the expression procedure was optimized including cultivated temperature, optional induction concentration and time of IPTG. The result indicated that the nucleocapsid protein can be expressed efficiently with 0.8mmol/L IPTG and 4 hour induction. The resulting Trx-N recombinant fusion protein was identified to be consisted of 34 kDa protein by SDS-PAGE and western blotting analysis. It indicated that the recombinant fusion protein could be used as antigen of diagnostic assay for detecting antibodies.