| 贾一琼1 邢益平1△ 王文娟1 李军1 王世霞2 卢山2 黄祖瑚1.外分泌型的乙型肝炎病毒核心抗原核酸疫苗的构建与体外表达[J].,2012,12(16):3090-3092 |
| 外分泌型的乙型肝炎病毒核心抗原核酸疫苗的构建与体外表达 |
| Construction and in Vitro Expression of Genetic Vaccine Encoding HepatitisB Virus Core Antigen Fused to Tissue Plasminogen Activator SignalSequence |
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| DOI： |
| 中文关键词: 乙型肝炎病毒核心抗原 组织纤溶酶原激活剂 质粒构建 核酸疫苗 |
| 英文关键词: Hepatitis B virus core antigen tPA leader Plasmid construction DNA vaccine |
| 基金项目:2011 年江苏省医学创新团队序号21 |
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| 中文摘要: |
| 目的：构建含有组织型纤溶酶原激活剂(tPA)信号肽的乙型肝炎病毒核心抗原(HBcAg)核酸疫苗。方法将HBcAg 基因用
PCR 方法扩增并分别引入相应的限制性内切酶酶切位点，然后克隆入pJW4303 载体中获得相应的核酸疫苗，经筛选鉴定后，用上
述核酸疫苗与野生型HBcAg 核酸疫苗及空载体质粒pJW4303 分别用脂质体瞬时转染293T 细胞，应用蛋白印迹法检测核心抗原
的表达。结果成功构建以pJW4303 为载体的含tPA 信号肽的HBcAg 核酸疫苗，体外表达证实含tPA 信号肽的HBcAg 在293T
细胞胞内和胞外均能表达，含tPA 信号肽的HBcAg 核酸疫苗的核心抗原表达水平较高。结论以pJW4303 为载体的含tPA 信号
肽的HBcAg 核酸疫苗能够将细胞内的HBcAg 分泌到细胞外，为进一步研究其免疫原性打下了基础。 |
| 英文摘要: |
| Objective: To construct and express nucleic acid vaccine encoding hepatitis B virus core antigen(HBcAg) fused to tissue
plasminogen activator signal sequence(tPA) in vitro. Methods: Restriction sites were introduced to HBcAg cDNA using polymerase chain
reaction(PCR). The gene product was cloned to pJW4303 to construct nucleic acid vaccine and all constructs were identified by sequencing,
and 293T cells were transiently transfected with pJW4303/HBc/tPA, pJW4303/HBc and pJW4303 blank vector. The HBcAg protein
was measured by western blot. Results: pJW4303/HBc/tPA nucleic acid vaccine was constructed successfully and expressed HBcAg in
lysates and supernatants in 293t cells in vitro. Expression level of pJW4303/HBcAg/tPA was higher than that of pJW4303/HBc. Conclusion:
HbcAg encoded by nucleic acid vaccine pJW4303/HBc/tPA and the expressed product in 293t cells could be secreted out of the cells.
This research is the foundation for the further study of the pJW4303/HBc/tPA nucleic acid vaccine. |
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