人乳头瘤病毒16型L1蛋白的克隆及表达

采用PCR技术从宫颈癌组织中扩增人乳头瘤病毒16型(Human papillomavirus type16,HPV16)L1全长基因片段,目的片段克隆到pMD18T载体后经酶切鉴定及测序确认。构建重组原核表达质粒pGEX4T1-L1,转化大肠杆菌E.coliBL21,IPTG诱导表达出以非可溶性蛋白形式存在的表达蛋白,该重组蛋白的表达量占菌体总蛋白的17%,免疫印迹检测表明,表达蛋白与宫颈癌病人血清出现特异性反应。成功构建了重组原核表达质粒pGEX4T1-L1,并且在原核细胞中得到表达,为进一步研究L1蛋白的免疫学活性及疫苗的研发奠定了基础。

Cloning and expression of L1 protein of human papillomavirus type 16 (HPV16)

The full-length of L1 gene of HPV16 was amplified from cervical cancer tissue by PCR and inserted into pMD18T vector by T-A cloning.The cloned vector was confirmed by enzymes digestion and sequencing.The recombinant expression plasmid pGEX4T1-L1 was transformed into E.coli BL21 and induced by IPTG to express the target protein which was in a form of inclusion body.The target protein accounted for approximately 17% of total bacterial proteins.It could react with the serum of patients suffered from cervical cancer by Western blot analysis.The result showed that the recombinant prokaryotic expression vector pGEX4T1-L1 was successfully constructed and fusion protein was expressed in E.coli.It provided a basis for the investigation on the immune responses of L1 protein and potential development of vaccine.