应用PCR技术克隆人乳头瘤病毒16型E6基因

CLONING HPV16 E6 GENE WITH POLYMERASE CHAIN REACTION

为进一步研究与宫颈癌发生密切相关的人乳头瘤病毒16型E6基因的转化作用,我们运用PCR技术扩增HPV16 E6 DNA,以pUC—19为载体,大肠杆菌了JM103为宿主菌,组建了质粒pRCE6经限制性核酸内切酶和Southern转印杂交证实,其插入片段约为0.5Kb,含有全部HPV16 E6序列。该质粒的组建为检测HPV感染中mRNA转录提供了特异性的早期基因探针。同时为进一步研究HPV16 E6基因的转化作用及转化蛋白打下基础。

The human papillomavirus type 16 (HPV16) may play some important roles in the onco-genesis of cervical carcinoma. In order to investigate the transformation of HPV16 E6 gene, we amplified the HPV16 E6 DNA with PCR. Using pUC-19 as the vector, E. coli JM103 strain as the host cell, plasmid pRCES was constructed. Confirmed by REanalysis and Southern blot hybridization, the inserted band of pRCE6 was about 0. 5Kb containing the total E6 sequence. The pRCE6 could be used as the specific probe of HPV16E6 region for detecting viral mRNA in HPV16 infected cells. Meanwhile it was a basework for studying the transformation and transforming proteins of HPV16.