汉坦病毒H8205部分核壳蛋白基因在E.coli中的表达  

Expression of theS genome segment of HantavirusH8205 and Application of the Expressed Polypeptide for Diagnosis of HFRS

作　　者：沈昌贤[1] 李钟铎[1] 杨瑞馥[1] 祝庆余[1] 

机构地区：[1]军事医学科学院微生物流行病研究所

出　　处：《中国病毒学》1999年第2期110-115,共6页Virologica Sinica

摘　　要：根据汉坦病毒Ｈ８２０５株ＮＰ基因的序列，设计一对引物，扩增ＮＰ前２９２个氨基酸多肽基因片段，克隆于表达载体ｐＧＥＸ３Ｘ，与载体中２６ｋＤ的谷胱苷肽巯基转移酶（ＧＳＴ）融合表达。ＳＤＳＰＡＧＥ显示表达产物（ＧＳＴＮＰ）主要以包涵体形式存在。Ｗｅｓｔｅｒｎｂｌｏｔｉｎｇ表明此融合蛋白有抗原性。包涵体经分离、洗涤、溶解后，Ｓｅｐｈａｒｏｓｅ６Ｂ层析纯化，用此融合蛋白作抗原，进行ＥＬＩＳＡ法检测临床ＨＦＲＳ病人标本的ＩｇＧ和ＩｇＭ，有很好的特异性和敏感性。有生物活性的汉坦病毒Ｈ８２０５ＮＰ的体外表达成功，为汉坦病毒基因工程抗原的大量制备奠定了基础，也为汉坦病毒的临床检测和流行病学调查提供了一种廉价、安全、可靠的抗原。The S genome segment encoding the nucleocapsid protein (truncated) of Hantavirus H8205 was amplified by PCR and cloned into the expression vector pGEX 3X and expressed in E coli. SDS PAGE showed that the expressed fusion protein (GST NP) was mainly in the form of inclusion body. The expressed protein was used as diagnostic antigen in solid phase enzyme immunoassay. The assay was used to detect IgG and IgM antibody in sera of HFRS patients originated from different geographic regions of China. The results revealed highly sensitive and specific. The successful expression and application of recombinant nucleocapsid protein of Hantavirus provides cheap, safe and sensitive antigen for the diagnosis of HFRS in China.

关 键 词：汉坦病毒 核衣壳蛋白 ELISA 

分 类 号：R373.32[医药卫生—病原生物学] Q939.4[医药卫生—基础医学]