Expression of the cp gene and Antiserum Preparation of Wisteria  Vein Mosaic Virus

LIANG Wen-xing; SONG Li-min; HUANG Jin-guang; TIAN Guo-zhong; LI Huai-fang; FAN Zai-feng

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June 2004

LIANG Wen-xing, SONG Li-min, HUANG Jin-guang, TIAN Guo-zhong, LI Huai-fang and FAN Zai-feng. Expression of the cp gene and Antiserum Preparation of Wisteria Vein Mosaic Virus[J]. Virologica Sinica, 2004, 19(3): 281-284.

Citation: LIANG Wen-xing, SONG Li-min, HUANG Jin-guang, TIAN Guo-zhong, LI Huai-fang, FAN Zai-feng. Expression of the cp gene and Antiserum Preparation of Wisteria Vein Mosaic Virus .VIROLOGICA SINICA, 2004, 19(3) : 281-284.

紫藤脉花叶病毒cp基因在大肠杆菌中的表达及抗血清的制备*

梁文星 ¹ ,
宋丽敏 ¹ ,
黄金光 ¹ ,
田国忠 ² ,
李怀方 ¹ ,
范在丰 ^1*,,

1.
1. 中国农业大学植物病理系，北京100094
2.
中国林业科学研究院森林生态环境与保护研究所，北京100091

通讯作者： 范在丰,

摘要

采用RT-PCR方法自紫藤脉花叶病毒北京分离物（WVMV-BJ）的基因组中分离出其CP基因，连接到原核表达载体pET22b(+)上。获得的重组子pET-WVMVCP转化大肠杆菌BL21（DE3）后，用IPTG进行诱导表达。SDS-PAGE和Western blot分析表明，cp基因在大肠杆菌中获得了高效表达，融合蛋白分子量约为34.4 kDa。将融合蛋白纯化后免疫兔子，获得了特异性较高的抗血清。微量免疫沉淀法测定该抗血清的效价为1/1024，酶联法 (enzyme-linked immunosorbant assay，ELISA)测定的效价为1/8192。
- 紫藤脉花叶病毒
- , 外壳蛋白
- , 原核表达
- , 抗血清
- , ELISA

Expression of the cp gene and Antiserum Preparation of Wisteria Vein Mosaic Virus

1.
1.Department of Plant Pathology, China Agricultural University, Beijing 100094, China
2.
Research Institute of Forest Ecology, Environment and Protection, China Academy of Forestry, Beijing 100091, China

Corresponding author: FAN Zai-feng,

Abstract

The coat protein (CP) gene of the Beijing isolate of Wisteria vein mosaic virus (WVMV-BJ) was amplified by RT-PCR, and ligated to the expression vector pET22b(+). The recombinant plasmid pET-WVMVCP was transformed into E. coli BL21(DE3) and then induced by IPTG. It was shon that the CP gene was highly expressed by SDS-PAGE and Western blot analysis. The molecular weight of the recombinant protein was about 34.4kDa. Antiserum with high specificity was produced after the rabbit was immunized with purified recombinant protein, and the titer was determined to be 1/1024 by micro-precipitation, or 1/8192 by antigen coating plate- ELISA(ACP-ELISA).
- Wisteria vein mosaic virus
- , CP
- , Prokaryotic expression
- , Antiserum
- , ELISA

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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 the cp gene and Antiserum Preparation of Wisteria Vein Mosaic Virus

Corresponding author: FAN Zai-feng,

1. 1.Department of Plant Pathology, China Agricultural University, Beijing 100094, China
2. Research Institute of Forest Ecology, Environment and Protection, China Academy of Forestry, Beijing 100091, China

Abstract: The coat protein (CP) gene of the Beijing isolate of Wisteria vein mosaic virus (WVMV-BJ) was amplified by RT-PCR, and ligated to the expression vector pET22b(+). The recombinant plasmid pET-WVMVCP was transformed into E. coli BL21(DE3) and then induced by IPTG. It was shon that the CP gene was highly expressed by SDS-PAGE and Western blot analysis. The molecular weight of the recombinant protein was about 34.4kDa. Antiserum with high specificity was produced after the rabbit was immunized with purified recombinant protein, and the titer was determined to be 1/1024 by micro-precipitation, or 1/8192 by antigen coating plate- ELISA(ACP-ELISA).