论文部分内容阅读
在狂犬病的临床诊断和基础研究中都迫切需要大量纯度高且价廉的狂犬病毒糖蛋白抗原。本文应用带有Ｈｉｓ６尾的ｐＥＴ原核表达系统对狂犬病毒（ＲＶ）ａＧ株的糖蛋白（ＧＰ）进行表达和纯化。构建的融合表达载体ｐＥＴ－ａＧ１和ｐＥＴ－ａＧ２（－５７ｂｐ）分别含有ＲＶａＧ株ＧＰ基因的全序列及删除了为ＧＰ信号肽编码的５８个碱基的序列。用ＳＤＳ聚丙烯酰胺凝胶电泳、免疫印迹、间接ＥＬＩＳＡ检测都证明表达产物为ＲＶＧＰ，且位于菌体中的包涵体内。经固定化金属螯合层析（ＩＭＡＣ）提纯，ｐＥＴ－ａＧ１表达产物有较高的特异性和纯度，可用作测定ＲＶＧＰ抗体的免疫诊断试剂。
In the clinical diagnosis of rabies and basic research are urgently needed a large number of high purity and cheap rabies virus glycoprotein antigen. In this study, the glycoprotein (GP) of aG strain of rabies virus (RV) was expressed and purified by using the pET prokaryotic expression system with His6 tail. The constructed fusion expression vectors pET-aG1 and pET-aG2 (-57bp) contained the complete sequence of the GP gene of RVaG strain and deleted the 58 base sequence encoded by GP signal peptide, respectively. SDS-polyacrylamide gel electrophoresis, Western blot, indirect ELISA test showed that the expression product RVGP, and located in the body of the inclusion body. After purification by immobilized metal chelation chromatography (IMAC), the pET-aG1 expression product has high specificity and purity and can be used as an immunodiagnostic reagent for the determination of RVGP antibodies.