摘要
目的:构建I型单纯疱疹病毒型特异性包膜糖蛋白G基因片段的表达载体,进行原核表达,并纯化目的蛋白。方法:用PCR法扩增HSV1-gG强抗原决定簇的基因片段,将该段基因克隆于PBV221表达载体,转化大肠杆菌DH5α,温控诱导目的蛋白表达,表达产物以包涵体形式存在,用SDS-PAGE和Westernblot鉴定目的蛋白,经离子交换纯化获得较纯的目的蛋白。结果:获得了重组的原核表达载体及相对分子质量为14.7kD的重组蛋白,并纯化获得了纯度大于90%的目的蛋白抗原。结论:成功克隆和表达了高纯度的HSV1-gG蛋白。
Objective: To construct the prokaryotic expression vector of specific-glycoprotein G gene fragment of herpes simplex virus type 1 (HSV-1), and express and purify the target protein. Methods: The HSV-1 glycoprotein G gene fragment containing dominant antigenic determinants was amplified by PCR and cloned into the expressive vector PBV221, then transferred into E. Coli DH5α. The recombinant was induced by increasing temperature to express the target protein, which existed in the form of inclusion. The protein was determinated by SDS-PAGE and Western blot, and purified through ion exchange to obtain the purer target protein. Results: The prokaryotic expression vector was constructed successfully and expressed the recombination protein of 14.7kD, which reached a purity over 90% by purification and showed good antigenicity by Western blot. Conclusion: HSV1-gG gene fragment is cloned and expressed successfully, which lay a foundation of developing good reagents for HSV1 immunoassay.
出处
《山东大学学报（医学版）》
CAS
北大核心
2005年第6期473-477,共5页
Journal of Shandong University:Health Sciences
基金
山东省重点攻关项目(2004GG2202150)
山东省优秀中青年科学家奖励基金
