Expression of Domain III of the DENV-2 E Protein and Preparation of Anti-His-D2EIII Polyclonal Antibody

ZOU Gang; QING Min; CAI Quan-xin; YUAN Zhi-ming

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December 2006

ZOU Gang, QING Min, CAI Quan-xin and YUAN Zhi-ming. Expression of Domain III of the DENV-2 E Protein and Preparation of Anti-His-D2EIII Polyclonal Antibody[J]. Virologica Sinica, 2006, 21(6): 619-621.

Citation: ZOU Gang, QING Min, CAI Quan-xin, YUAN Zhi-ming. Expression of Domain III of the DENV-2 E Protein and Preparation of Anti-His-D2EIII Polyclonal Antibody .VIROLOGICA SINICA, 2006, 21(6) : 619-621.

登革2型病毒E蛋白结构域III的表达及多克隆抗体制备

1.
中国科学院武汉病毒研究所，湖北武汉 430071

通讯作者： 袁志明,

摘要
- 登革2型病毒
- , E蛋白
- , 结构域III
- , 亲和层析
- , 多克隆抗体

Expression of Domain III of the DENV-2 E Protein and Preparation of Anti-His-D2EIII Polyclonal Antibody

Corresponding author: YUAN Zhi-ming,

Abstract

The gene fragment coding for amino acids 281 to 395 of the E protein of DENV-2 （New Guinea C strain） was amplified by PCR, comprising Domain III (amino acids 295 to 395) of the E protein. The fragment was cloned into pMD18-T vector and subcloned to expression vector pET-28a and pMAL-c2X. The recombinant plasmid pET-28a-D2EIII was transformed into E.coli BL21(DE3) and the pMAL-c2X-D2EIII was transformed into E.coli TB1. The induced recombinant proteins were purified by His-tag and MBP-tag affinity chromatography, respectively. The purified protein His-D2EIII was used to immunize rabbit three times at two-week intervals, the immunized rabbit produced high titer anti-His-D2EIII polyclonal antibody. The result of western blot indicated that the expressed fusion protein could react with the polyclonal antibody against Domain III of E protein.
- DENV-2
- , E protein
- , Domain III
- , Affinity chromatography
- , Polyclonal antibody

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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

Expression of Domain III of the DENV-2 E Protein and Preparation of Anti-His-D2EIII Polyclonal Antibody

Corresponding author: YUAN Zhi-ming,

Abstract: The gene fragment coding for amino acids 281 to 395 of the E protein of DENV-2 （New Guinea C strain） was amplified by PCR, comprising Domain III (amino acids 295 to 395) of the E protein. The fragment was cloned into pMD18-T vector and subcloned to expression vector pET-28a and pMAL-c2X. The recombinant plasmid pET-28a-D2EIII was transformed into E.coli BL21(DE3) and the pMAL-c2X-D2EIII was transformed into E.coli TB1. The induced recombinant proteins were purified by His-tag and MBP-tag affinity chromatography, respectively. The purified protein His-D2EIII was used to immunize rabbit three times at two-week intervals, the immunized rabbit produced high titer anti-His-D2EIII polyclonal antibody. The result of western blot indicated that the expressed fusion protein could react with the polyclonal antibody against Domain III of E protein.