伪狂犬病病毒鄂A株囊膜蛋白gD基因在杆状病毒载体系统中的表达研究  

Expression of the Envelope Glycoprotein D Gene of Pseudorabies Virus Ea Strain

作　　者：陈新华[1] 杨林[2] 龙綮新[2] 王章 陈曲侯[3] 洪文洲[4] 陈焕春[4] 

机构地区：[1]国家海洋局第三海洋研究所海洋生物工程重点实验室,厦门361005 [2]中山大学生物防治国家重点实验室,广州510275 [3]华中师范大学昆虫学研究所,武汉430079 [4]华中农业大学畜牧兽医学院,武汉430070

出　　处：《生物化学与生物物理进展》2002年第3期415-419,共5页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金 (3 9670 5 5 );瑞典青年基金 (IFS :B/2 987 1)资助项目

摘　　要：克隆了伪狂犬病病毒鄂A株囊膜蛋白gD基因并进行了序列分析 ,与国际标准毒株Rice株相比 ,两者之间核苷酸序列同源性为 98% ,推导氨基酸序列同源性为 97% .利用杆状病毒GST融合载体系统对其进行了高效表达 ,SDS 聚丙烯酰胺凝胶电泳及蛋白质印迹均证明了表达产物为 71ku的GST gD融合蛋白 ,其产量占细胞不溶蛋白量的 2 0 %左右 .以GST gD融合蛋白作为抗原进行小鼠免疫保护实验 ,结果表明表达的GST gD融合蛋白具有较好的免疫原性 ,不仅能诱导小鼠产生血清抗体 (1∶12 8) 。The envelope glycoprotein D gene of pseudorabies, virus Ea strain was cloned by PCR technique. Sequence analysis displayed 98% nucleotide sequence homology and 97% deduced amino acid sequence homology between the cloned gD gene and PRV Rice strain gD gene. Then gD gene was expressed highly in the baculovirus GST fusion vector system. Both SDS-PAGE and Western-blot verified that the expression product was GST-gD fusion protein with the. molecular mass of 71 ku. Expressed GST-gD fusion protein accounted for about 20% of total cellular protein. The protective immune assay was performed in mice by using GST-gD fusion protein as antigen. The result, showed that all mice immunized by GST-gD could produce a certain level of antibody against PRV gD (antibody titer: 1: 128), and the mice in immunized group could be partly protected against challenge infection of highly virulent PRV Su strain at 2 x 10(5) PFU per mice. A good foundation has been laid down for developing PRV genetically-engineered subunit vaccine.

关 键 词：伪狂犬病病毒 鄂A株 囊膜蛋白gD基因 杆状病毒载体系统 表达 免疫原性 

分 类 号：S852.659.1[农业科学—基础兽医学]