兔出血症病毒NJ85株衣壳蛋白基因在原核系统中的高效表达  

High Expression of the Capsid Protein Gene of the Chinese Early Isolate NJ85 of Rabbit Hemorrhagic Disease Virus in Escherichia coli

作　　者：王永山[1] 陆承平[1] 周宗安 

机构地区：[1]南京农业大学动物医学院,南京210095 [2]华东医学生物技术研究所,南京210002

出　　处：《农业生物技术学报》2004年第4期408-411,共4页Journal of Agricultural Biotechnology

基　　金：RHDV NJ85毒株VP60基因序列已在GenBank注册(GenBank accession AY269825)。

摘　　要：从兔出血症病毒(Rabbit hemorrhagic disease virus,RHDV)中国早期流行株NJ85中成功地克隆出衣壳蛋白(VP60)基因。序列分析表明该基因长度为1 740 nt,编码579 aa,与已报道的另2个RHDV中国毒株WX84和TP 的VP60基因的同源性分别为92.7%和97.2%。构建了NJ85株VP60基因原核表达质粒pET-VP60,用SDS-PAGE分析,重组VP60蛋白的表达量占菌体总蛋白的40%,分子量约62 kD,在菌体内以包涵体形式存在。免疫印迹实验表明,重组VP60蛋白与RHDV抗体反应形成1条带,证明它具有抗原性。The capsid protein (VP60 ) gene of Rabbit hemorrhagic disease virus (RHDV) isolate NJ85 was amplified and sequenced. The VP60 gene of NJ85 was in size of 1 740 nt and encoding 579 aa. Comparison with the other Chinese field isolates WX84 and TP , the homology of nucleotide sequence was 92.7% and 97.2% respectively. The expression plasmid pET-VP60 contained the full-length VP60 gene from NJ85 was constructed and highly expressed under the induction with IPTG. The proportion of the recombinant VP60 protein reached to 40 % of the total bacterial proteins under the optimized inductive condition. SDS-PAGE analysis showed that the molecular weight of the recombinant VP60 protein was approximate to 62 kD, and it formed inclusion body in Escherichia coli. The specific reaction of the recombinant VP60 protein with antiserum of RHDV was confirmed by immunoblotting assay.

关 键 词：兔出血症病毒 衣壳蛋白基因 高效表达 原核系统 VP60 原核表达质粒 RHDV virus 基因长度 PAGE 免疫印迹 抗体反应 流行株 分析表 同源性 pET SDS 总蛋白 表达量 分子量 抗原性 中国 重组 克隆 体形 

分 类 号：Q78[生物学—分子生物学] S858.291[农业科学—临床兽医学]