汉滩病毒核蛋白羧基端多肽真核表达载体的构建及其表达产物的鉴定  

Construction of HV whole and partial S gene segments eukaryotic expression vectors and identification of expressed nucleocapsid protein

作　　者：李光玉[1] 白雪帆[1] 黄长形[1] 潘蕾[1] 赵玉玲[1] 陈红梅[1] 韦三华[1] 杨为松[1] 

机构地区：[1]第四军医大学唐都医院全军感染病防治中心,陕西西安710038

出　　处：《第四军医大学学报》2002年第3期243-245,共3页Journal of the Fourth Military Medical University

摘　　要：目的　探讨汉滩病毒 (HTNV)核蛋白羧基端多肽的抗原性 ,为建立分型检测方法奠定基础 .方法　设计并合成 S基因编码区及其 3 端 82 5 bp片段引物 ,用 PCR方法从PBV2 2 0 - S2 2原核质粒中扩增出目的片段 ,应用 TA克隆将其插入 pc DNA3.1/ V5 - His- TOPO载体中 ,并通过脂质体转染至 Vero细胞中进行瞬时表达 ,表达产物用间接免疫荧光法进行鉴定 .结果　成功构建了 pc DNA3.1- S及 pc DNA3.1- S- C真核表达载体 ,间接免疫荧光法可检测到真核表达质粒转染的 Vero细胞表达的核蛋白羧基端多肽 .结论　pc DNA3.1- S及 pc DNA3.1- S- C真核表达载体有较高的转染效率 ,能在宿主细胞中表达 。AIM To study the antigenicity of eukaryotic expressed products of HV S partial gene (502~1326 bp) and obtain some data for serotyping antigen. METHODS The eukaryotic expression vectors pcDNA3.1 S and pcDNA3.1 S C were constructed with pcDNA 3.1/V5 His TOPO TA Cloning Kit, then the recombinant expression plasmids were transfected into Vero cells in vitro . The transient expression products were analyzed by IFA. RESULTS It was provedthat the eukaryotic expression vectors pcDNA3.1 S and pcDNA3.1 S C were successfully constructed by restriction enzyme analysis and sequence analysis. IFA results showed that the expressed protein could be detected distinctly and peculiarly. CONCLUSION The truncated NP (aa155 to 429) could be considered as serotyping antigen.

关 键 词：汉滩病毒 核衣壳蛋白 多肽抗原 真核表达 载体构建 表达产物 鉴定 

分 类 号：R373.9[医药卫生—病原生物学]