猪戊型肝炎病毒swCH189株衣壳蛋白基因CP239片段原核表达条件的优化  

Optimization of Prokaryotic Expression for Capsid Protein Gene CP239 in the swCH189 Strain of Swine Hepatitis E Virus

作　　者：郝宝成[1,2] 梁剑平[1,3] 兰喜[2] 刑小勇[3] 项海涛[3] 温峰琴[3] 胡永浩[3] 柳纪省[2,3] 

机构地区：[1]中国农业科学院兰州畜牧与兽药研究所/新兽药重点开放实验室/甘肃省新兽药工程重点实验室,兰州730050 [2]中国农业科学院兰州兽医研究所/家畜疫病病原生物学国家重点实验室/农业部畜禽病毒学重点开放实验室/农业部草食动物疫病重点开放实验室,兰州730046 [3]甘肃农业大学动物医学院,兰州730070

出　　处：《湖北农业科学》2011年第15期3116-3119,共4页Hubei Agricultural Sciences

基　　金：中央级公益性科研院所基本科研业务费专项资金(BRF090402)

摘　　要：为了提高猪戊型肝炎病毒swCH189株衣壳蛋白基因CP239片段在大肠杆菌中的表达量,研究了载体、温度、转速、诱导时间以及诱导剂IPTG浓度等不同条件对衣壳蛋白基因CP239片段融合蛋白表达量的影响。结果表明,用LB培养基于37℃培养3.5 h后,采用终浓度为0.3 mmol/L的IPTG在37℃、200r/min诱导培养4 h,pET32a-CP239融合蛋白表达量最大;SDS-PAGE检测结果表明pET32a-CP239融合蛋白的分子质量与预期大小一致,约为45.3 ku;Western blotting结果表明,pET32a-CP239融合蛋白可以与抗-HEV阳性血清发生特异性反应,并具有良好的反应原性,说明衣壳蛋白基因CP239片段蛋白得到正确表达。To improve the expression quantity of capsid proteins genes CP239 fragment of swine hepatitis E virus swCH189 strains in Escherichia coli.,the influence of vector,temperature,rotating speed,inducting time and IPTG concentration on the expression quantity was analysed.The results showed that the fusion protein expression amount of pET32a-CP239 was the largest after cultivated 3.5 h with LB at 37 ℃ and 4 h inducting cultured with 0.3 mmol/L IPTG at 37 ℃,200 r/min.The molecular weight was determined as 45.3 ku by SDS-PAGE electrophoresis which was agreement with prediction.The Western blotting detection showed a specific reaction of pET32a-CP239 fusion protein for Anti-HEV positive serum and with well reactionogenicity.It indicated that the capsid protein gene CP239 fragments was properly expressed.

关 键 词：猪戊型肝炎病毒 衣壳蛋白 CP239片段 原核表达 

分 类 号：Q786[生物学—分子生物学]