靶向乳头瘤病毒18型E6基因的shRNA表达载体的构建与鉴定  

The construction and verification of a vector expressing small hairpin RNA targeting Human Papillomavirus 18 E6

作　　者：王雪莲[1] 王冬冬[2] 安春丽[1] 张丽[3] 李唯[4] 肖纯凌[5] 

机构地区：[1]中国医科大学基础医学院病原生物学教研室,辽宁沈阳110001 [2]中国医科大学附属盛京医院妇产科,辽宁沈阳110003 [3]中国医科大学附属第一医院皮肤科,辽宁沈阳110001 [4]中国医科大学附属第一医院儿科,辽宁沈阳110001 [5]沈阳医学院病原生物学教研室,辽宁沈阳110034

出　　处：《中国病原生物学杂志》2010年第1期1-5,共5页Journal of Pathogen Biology

基　　金：国家自然科学基金项目(No.30670916);辽宁省科学技术项目(No.2007225005-22)

摘　　要：目的以乳头瘤病毒(human papillomavirus,HPV)18型E6基因为靶标,构建小发夹状RNA(small hairpinRNA,shRNA)表达载体。方法人工方法合成针对靶序列的两条寡核苷酸链,两端加入酶切位点。寡核苷酸链退火后与酶切的线性化表达载体连接。连接产物转化大肠埃希菌,卡那霉素筛选阳性克隆。结果重组质粒酶切片段约为400 bp,与插入片段大小一致。DNA序列分析证实插入片段序列与预期一致。结论成功构建HPV shRNA表达载体pHPV1、pHPV2。Objective To construct a vector targeting small hairpin RNA expressing Human Papillomavirus （HPV） 18 E6. Methods Oligonueleotides with enzyme cleavage sites were synthesized, annealed, and then ligated with the plasmid pGenesil cleaved with enzymes. The product was transformed into E. coli DH5a. Positive clones were selected. Results The size of cleaved fragments of restriction enzymes was in accordance with the size of the inserted fragments. DNA sequencing also showed that the sequence of the inserted fragments was the same as that expected. Conclusion pHPV1 and pHPV2 vectors expressing shRNA were successfully constructed.

关 键 词：乳头瘤病毒 小发夹状RNA E6 

分 类 号：R373.9[医药卫生—病原生物学]