丙型肝炎病毒核心区多肽的抗原表位分析和原核表达

Study on the Epitopes of the Polypeptide in the Core Region of Hepatitis C Virus and its Prokaryotic Expression

目的分析HCV核心区多肽的抗原表位,选择具有优势抗原表位的多肽用原核表达载体表达,并获得该表达载体的稳定表达菌株。方法根据软件分析选取带有优势抗原表位的HCV核心区多肽,找出对应DNA序列,逆转录PCR扩增目的基因,克隆到原核表达载体中进行诱导表达。表达的目的蛋白用His Bind Columns纯化,用ELISA法检测其免疫活性。结果获得了序列正确的HCV-core基因和表达良好的该基因原核表达载体pET-28a-core,该表达载体表达的目的蛋白可被很好地纯化并具有良好的HCV抗原活性。结论成功地重组了HCV核心区蛋白,获得了HCV抗原性良好的HCV核心区抗原。

Objective To analyze the epitopes of the polypeptide in the core region of hepatitis C virus （HCV） and to select the polypeptide fragments with preponderant antigenic determinants to express in prokaryotic expression vector to obtain the strain with stable expression of the vector. Methods The polypeptide fragments in the HCV core region comprising the pre- ponderant antigenic determinants were selected by software analysis, and the corresponding DNA sequences were searched and the target genes were amplified by reverse transcription PCR. Then the gene fragment was cloned into the prokaryotic expression vector and its expression was induced. The expressed target protein was purified by His Bind Columns, and its immunocompetence was determined by ELISA. Results The HCV - core gene with right sequences and its prokaryotic expression vector with high efficiency, pET- 28a- core, were obtained. The expressed target protein could be well purified and had outstanding HCV antigenicity. Conclusions HCV core protein is successfully recomposed, and the HCV core protein with excellent HCV anti- genicity is obtained.