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目的构建含乙肝核心C基因的真核表达载体pcDNA3.1-HBcNheI,探索其作为DNA疫苗载体的可能性。方法采用分子克隆技术在原核表达质粒pTrc-core的HBcAgel-loop处添加NheI酶切位点,双酶切此重组载体pTrc-coreNheI及真核表达载体pcDNA3.1,产物经纯化、T4DNA连接酶连接,连接产物转化大肠杆菌DH5α,筛选抗Amp+克隆并提取质粒进行酶切及PCR鉴定。结果成功构建了真核表达质粒pcDNA3.1-HBcNheI。结论该重组质粒可作为治疗性及预防性疫苗的候选载体,为深入研究预防性及治疗性疫苗奠定了基础。
Objective To construct the eukaryotic expression vector pcDNA3.1-HBcNheI containing hepatitis C core C gene and explore its potential as a DNA vaccine vector. Methods The recombinant plasmid pTrc-coreNheI and its eukaryotic expression vector pcDNA3.1 were double digested with HBeAg-loop at the HBcAgel-loop of prokaryotic expression plasmid pTrc-core by molecular cloning. The product was purified and ligated with T4 DNA ligase , The ligation product was transformed into E. coli DH5α, anti-Amp + clones were screened and plasmids were extracted for enzyme digestion and PCR identification. Results The eukaryotic expression plasmid pcDNA3.1-HBcNheI was successfully constructed. Conclusion The recombinant plasmid can be used as a candidate vector for therapeutic and prophylactic vaccine, which lays the foundation for further study on preventive and therapeutic vaccines.