【摘要】 对构建HPV16 E6基因的原核表达质粒pET28a(+)-HPV16 E6进行研究.从宫颈癌细胞株CaSki细胞中提取的的DNA基因组为模板,设计一对特异性引物,用PCR法扩增出HPV16E6基因片段.HPV16E6基因片段和质粒pET28a(+)经NcoⅠ和HindⅢ双酶切后用T4 DNA连接酶连接.构建的重组质粒pET28a(+)-HPV16 E6转化大肠杆菌DH5α,经PCR鉴定和酶切鉴定筛选出阳性克隆,将阳性克隆的质粒转化到大肠杆菌BL21 Star-DE3 Plyss中,构建了HPV16 E6基因原核表达质粒pET28a(+)-HPV16E6.HPV16E6基因原核表达质粒pET28a(+)-HPV16 E6的构建为HPV16 E6重组蛋白的制备和为宫颈组织HPV16型感染的早期诊断和相关治疗性疫苗的研制奠定了物质基础.更多还原

【Abstract】 The objective of this paper is to build HPV16 E6 gene expression plasmid pET28a(+)-HPV 16 E6.The genomic DNA extracted from CaSki cells are employed as a template,a specific pair of primers is designed,HPV16E6 gene fragment is amplified by PCR.HPV16E6 gene fragment and plasmid pET28a(+) digested by NcoⅠand Hind Ⅲ are connected by T4 DNA ligase.Results show that the construction of recombinant plasmid pET28a(+)-HPV16 E6 is transferred into E.coli DH5α,the positive clones are screened by PCR and enzyme digestion,which are transferred into E.coli BL21 DE3 Star Plyss,HPV16E6 gene expression plasmid pET28a(+)HPV16E6 are built successfully.It is concluded that HPV16 E6 gene expression plasmid pET28a(+)-HPV 16 E6 is ready to prepare HPV16E6,which provides a foundation of the early diagnosis for HPV16 in cervical tissue and therapeutic vaccine about HPV16.更多还原