人乳头瘤病毒16型(新疆株)E7基因的克隆与突变研究  

Cloning and Mutation of Human Papillomavirus Type 16(Xinjiang strain)E7 Gene

作　　者：张茂祥[1] 张富春[1] 蹇晓红[1] 马正海[1] 王芸[1] 张馨玉[1] 王宾[1] 

机构地区：[1]新疆大学生命科学与技术学院分子生物学重点实验室,新疆生物资源基因工程重点实验室,新疆乌鲁木齐830046

出　　处：《生物技术》2003年第6期4-6,共3页Biotechnology

基　　金：国家自然科学基金项目 ("人乳头瘤病毒 1 6型E7核酸疫苗的研究" ;39960 0 79)资助

摘　　要：目的 :克隆分析人乳头瘤病毒 16型 (HPV16 )新疆株的E7基因 ;并对E7基因进行突变改造 ,以比较野生型与突变型HPV16E7基因的功能。方法 :根据从中国新疆维吾尔族妇女宫颈癌活检组织标本中提取的DNA ,进行PCR扩增获得HPV16E7基因 ,然后分别将其克隆到pMD18-T载体上进行DNA序列分析。根据HPV16E7基因的特点 ,分别设计点突变引物 ,用PCR的方法进行HPV16E7基因的点突变。结果 :PCR检测显示扩增出HPV16 (新疆株 )E7基因 ;测序结果表明HPV16 -XJ的E7基因全长2 97bp ,与德国标准株一致 ;利用设计突变位点的引物经PCR扩增 ,经序列测定后 ,分别得到了第 70、172、2 71位碱基突变的HPV16E7基因 ;分别构建了野生型与单、双、三点突变的重组质粒pMD18-T -HPV16E7。结论 :人乳头瘤病毒 16型 (新疆株 )E7基因结构与德国标准株相同。HPV16E7基因多点突变的改造 ,为探索HPV16E7基因功能的变化和开展疫苗研究奠定了理论基础。Objective:Cloning and mutations of HPV16 E7 Gene isolated from some Chinese Uygur patients of cervical carcinoma in South Xinjiang.Methods:HPV16 E7 gene was amplified by PCR with the template DNA,which was extracted from the cervical carcinoma tissue.This PCR product was cloned into pMD18-T and subjected to DNA sequence analysis.In order to study HPV16E7 biological functions,three pair of primers with the desired mutations were used to generate the mutant clones for HPV16 E7 Gene.Results:The successful E7 PCR product was cloned and designated as the pMD18-T-HPV16 E7.The sequence analysis shows that the HPV16 E7 from our clone is identical to that E7 from German standard strain with 297bp in length.The mutations are introduced from original Ts to Gs at 70,172 and 271 nucleotides of HPV16 E7 resulting three amino acids changes at the 24th,58th,91th of HPV16E7 protein sequence.Conclusions:There is the same sequence observed between HPV16 E7 gene of Xinjiang strain and the German standard strain.The mutations introduced to HPV16E7 gene can be used to develop a vaccine for HPV infection and immunotherapeutic approaches in future.

关 键 词：人乳头瘤病毒 E7基因 克隆 突变 

分 类 号：R373[医药卫生—病原生物学] Q939.4[医药卫生—基础医学]