Cloning and Expression of E Gene of Japanese Encephalitis Virus and Preliminary application

ZHANG Xue-han; HE Kong-wang; GUO Rong-li; NI Yan-fang; WANG Fang; YU Zheng-yu

PDF
Cite
Share

facebook

twitter

google

linkedin

October 2005

ZHANG Xue-han, HE Kong-wang, GUO Rong-li, NI Yan-fang, WANG Fang and YU Zheng-yu. Cloning and Expression of E Gene of Japanese Encephalitis Virus and Preliminary application[J]. Virologica Sinica, 2005, 20(5): 494-497.

Citation: ZHANG Xue-han, HE Kong-wang, GUO Rong-li, NI Yan-fang, WANG Fang, YU Zheng-yu. Cloning and Expression of E Gene of Japanese Encephalitis Virus and Preliminary application .VIROLOGICA SINICA, 2005, 20(5) : 494-497.

流行性乙型脑炎病毒E基因的克隆、表达及初步应用

1.
江苏省农业科学院兽医研究所农业部畜禽疫病诊断重点开放实验室江苏南京

摘要

参照GenBank中的日本乙型脑炎病毒(Japanese encephalitis virus,JEV)SA14-14株序列设计了一对特异性引物,用PCR方法从SA14-14扩增E基因全长,然后克隆到pMD18-T载体中,转化宿主菌DH5a,提取阳性克隆质粒进行双酶切鉴定,将目的片段定向克隆到pET32a(+)中,转化入BL21(DE3),经IPTG诱导可表达分子量约73ka的蛋白,Western blotting试验呈阳性,表明E基因得到表达。以纯化的表达产物为核心抗原,猪抗JEV血清为一抗,HRP标记羊抗猪IgG抗体为二抗建立间接ELISA方法,并初步检测了一些血清样品,结果提示表达的蛋白具有很好的应用开发价值。
- 乙型脑炎病毒
- , E基因
- , 表达
- , 间接ELISA

Cloning and Expression of E Gene of Japanese Encephalitis Virus and Preliminary application

Abstract

The whole cDNA of E gene was amplified by RT-PCR from JEV strain SA14-14,and cloned into the pMD18-T vector.The fragment was identified by restriction enzymes digestion with EcoR I and Xho I,cloned into the pET32a(+) vector.The recombinant plasmid was transformed into BL21 and the recombinant bacteria was induced by optimal concentration of IPTG.SDS-PAGE and Western blotting were performed to detect the E fusion protein.Our result indicates the molecular weight of E protein is of E protein 73kDa and its antigenicity is good.Indirect ELISA procedure has been developed with the purified E protein as antigen for detection of JEV.
- Japanese encephalitis virus（JEV）
- , E gene
- , Indirect
- , ELISA

References
Proportional views

PDF

Article Metrics

Article views(3996) PDF downloads(987) Cited by(0)

Proportional views

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 E Gene of Japanese Encephalitis Virus and Preliminary application

Abstract: The whole cDNA of E gene was amplified by RT-PCR from JEV strain SA14-14,and cloned into the pMD18-T vector.The fragment was identified by restriction enzymes digestion with EcoR I and Xho I,cloned into the pET32a(+) vector.The recombinant plasmid was transformed into BL21 and the recombinant bacteria was induced by optimal concentration of IPTG.SDS-PAGE and Western blotting were performed to detect the E fusion protein.Our result indicates the molecular weight of E protein is of E protein 73kDa and its antigenicity is good.Indirect ELISA procedure has been developed with the purified E protein as antigen for detection of JEV.