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目的 克隆ＨＧＶ Ｅ２ 基因并在原核细胞系统中表达ＨＧＶ Ｅ２ 蛋白。方法 从ＨＧＶＲＮＡ 阳性血浆中抽提病毒ＲＮＡ，利用半巢式ＲＴＰＣＲ 法扩增ＨＧＶ Ｅ２ 基因片段并进行序列分析。然后将该片段克隆到ｐＧＥＸ５ｘ１ 表达载体上，进行诱导表达。表达产物用抗ＨＧＶ 阳性血浆作Ｗｅｓｔｅｒｎ ｂｌｏｔ 活性鉴定。结果 获得ＨＧＶ Ｅ２ Ｎ 端长度为７７９ 个核苷酸的基因片段。该片段与美国株ＨＧＶ（Ｕ４４４０２） 、西非株ＧＢＶＣ（ Ｕ３６３８０） 、中国株ＨＧＶ（ Ｕ７５３５６） 的核苷酸序列同源性分别为８４ ％ 、８５ ％ 、９１ ％ ，推测的氨基酸序列同源性分别为８８ ％ 、９３ ％ 、９４ ％ 。表达产物为相对分子质量约５０ ×１０３的ＧＳＴＥ２ 融合蛋白，在细胞内形成包涵体。Ｗｅｓｔｅｒｎ ｂｌｏｔ 反应中在约５０ ×１０３ 处有显色条带。结论本研究成功地在原核细胞系统中表达了具有抗原性的ＨＧＶ Ｅ２ 蛋白，为进一步研究ＨＧＶ Ｅ２ 蛋白及Ｅ２ 抗体的生物学功能打下了基础。
Objective To clone HGV E2 gene and express HGV E2 protein in prokaryotic cell system. Methods The viral RNA was extracted from HGV RNA positive plasma and the HGV E2 gene fragment was amplified by semi-nested RT-PCR and sequenced. Then the fragment was cloned into pGEX 5x 1 expression vector for induction of expression. The expression product was identified by western blotting using anti-HGV positive plasma. Results The gene fragment of 779 nucleotides in length of N-terminal of HGV E2 was obtained. The nucleotide sequence of the fragment was 84%, 85%, 91% homologous to that of the American strain HGV (U44402), West African strain GBV C (U36380) and Chinese strain HGV (U75356) Sexuality was 88%, 93%, 94% respectively. The expression product was a GST-E2 fusion protein with a relative molecular mass of about 50 × 103, forming inclusion bodies in the cell. Western blot reaction at about 50 × 103 Department of color bands. Conclusion The present study successfully expressed the antigenic HGV E2 protein in prokaryotic cell system, which laid the foundation for further study of the biological function of E2 protein and HGV E2 antibody.