丙型肝炎病毒T细胞表位的筛选及其在大肠杆菌中的克隆与表达

目的：在丙型肝炎病毒(hepatitis C virus，HCV)不同变异株中，筛选高保守的能涵盖多种MHC限制类型的T细胞表位，并构建重组表位抗原的原核表达质粒。方法：检索文献并结合MHC限制型，从中遴选出功能明确的T细胞表位；利用Goldkey软件，建立HCV全长蛋白序列数据库；从中分别截取各个表位所在区段，对所选取的表位进行同源性比较；从中选取保守性强的部分表位，用化学法合成表位基因，插入原核表达载体pBVIL1中，构建多个融合表达质粒并在E．coli表达。结果：从文献中获得HCV免疫优势性T细胞表位18个，在由55条全长HCV序列组成的数据库中进行比较，获得了大量有关表位保守性的信息，并进行了统计分析，所构建的5个表达质粒pBVIL1—T1—T5经鉴定后，在大肠杆菌中均获得了高效表达。结论：本研究有助于深入认识HCV T细胞表位的特性，为表位的筛选提供了方法学依据，所选取的表位及原核表达的重组蛋白，为HCV治疗性疫苗的实验研究提供了候选抗原。

T-Cell antigenic epitopes of hepatitis C virus: selecting,cloning and expressing in E.coli

Objective:To select a series of HCV T cell epitopes highly conserved and multiple MHC supertype_restricted and to construct their expression plasmids.Methods:HCV T cell epitopes were selected from the published literature.By using the Goldkey software, a database containing whole HCV protein sequence was constructed and then all the selected epitopes were compared in this database. The highly conserved epitopes were selected and their genes were inserted into the expression vector pBVIL1 and expressed in E.coli .Results: Eighteen T cell epitopes were acquired from literature altogether. After comparing these epitopes in database including 55 HCV whole protein sequences,their homologies were clearly defined and the statistic analyses about all the epitopes were done.After testing, the epitope genes were proved to be inserted correctly into the vector and were expressed in E.coli highly.Conclusions:This work will do good to the awareness of the specialty of the T cell epitopes and will provide the methodology reference on the selection of the epitopes. It also paves the way of the study for the HCV therapeutic vaccination.