人乳头瘤病毒11型E6基因完整开放阅读框架的克隆和鉴定

Cloning and identification of complete open reading frame of E6 gene of human papillomavirus type 11

目的 为构建人乳头瘤病毒 1 1型 (HPV1 1 )DNA疫苗 ,从尖锐湿疣病变组织中克隆疫苗靶基因E6。方法 利用改良提取染色体外游离DNA的方法制备模板DNA ,采用聚合酶链反应(PCR)技术进行基因克隆。结果 从病变组织中扩增出了全长人乳头瘤病毒 1 1型E6基因完整开放阅读框架 ,并插入中介载体T easy中构建重组质粒 ,转化JM1 0 9扩增后经酶切、PCR及测序分析 ,证实克隆成功。结论 该实验的成功为下一步制备HPV1 1的治疗性疫苗奠定基础。

Objective To construct DNA vaccine against human papillomavirus type 11, target gene E6 was cloned from biopsy of condyloma tissues. Methods Episomal HPV DNA was extracted according to improved protocol and was subjected to PCR as template.Results The cloned gene was inserted into Teasy vector and the recombinant plasmid HPV11E6-Teasy was identified by endonucleases restriction digestion,PCR amplification and sequencing.A BLAST search confirmed that the cloned fragment contained the complete open reading frame of E6 gene of HPV11.Conclusion The target gene cloned from diseased tissue may provide a ready source for vaccine preparation.