人乳头瘤病毒11型E7与共刺激分子CD80嵌合DNA疫苗质粒的构建及鉴定

目的:构建人乳头瘤病毒11型(HPV ll)E7与共刺激分子CDS0嵌合DNA疫苗质粒。方法:从尖锐湿疣病变组织中提取DNA制备模板,采用聚合酶链反应(PCR)技术扩增HPV ll-E7基因,经双酶切后定向克隆到pcDNA3.1(+)中,获得pcDNA3.1(+)/HPV llE7,测序鉴定后,再用PCR方法扩增去除终止密码子的E7基因片段,按上述方法插入质粒pcD-NA3.1(+)/CD80中CD80上游,构建pcDNA3.1(+)/HPV llE7-CD80融合基因真核表达质粒。结果:去除终止子的E7基因片段成功正向插入CD80上游,双向测序分析显示序列无误,pcDNA3.1(+)/HPV llE7-CD80嵌合真核表达质粒构建成功。结论:HPV ll-E7/CD80嵌合DNA疫苗质粒构建成功,为研究该疫苗在尖锐湿疣防治中的作用奠定了基础。

Construction and identified of chimerical DNA vaccine plasmid of human papillomavirus type 11 E7 and CD80 co- stimulus gene

Objective:To construct chimerical DNA vaccine plasmid of human papillomavirus type 11(HPVll) E7 and CD80 co-stimulatory gene.Methods:DNA was extracted from biopsy of condyloma tissues and subjected to PCR as template.The gene fragment of HPV 1 I-E7 was amplified by polymerase chain reaction(PCR) and was cloned into the pcDNA3.1(+).The recombinant plasmid pcDNA3.1(+) / HPVIlE7 was identified by restriction end nuclease cleanses digestion and sequencing.The gene fragment of HPVl l-E7 without terminator was amplified by PCR and was cloned into the pcDNA3.1(+) /CD80 at the upstream of CD80.Results:The amplified fragment from diseased tissue contained HPV lI-E7 gene was inserted into the vector of pcDNA3.1(+)/CD80.Sequence in the constructed plasmids was correct.Conclusion:The chimerical DNA vaccine plasmid of pcDNA3.1(+)/HPVll E7-CDS0 is constructed successfully,which may be used for further research of the fusion protein vaccine for the preventive and treatment of genital wart.