| 人乳头状瘤病毒16型E6基因短发夹结构RNA慢病毒载体的构建及鉴定 |
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| 引用本文： | 尹倩,孙桂芹,庄艳艳,钟梅,苏桂栋.人乳头状瘤病毒16型E6基因短发夹结构RNA慢病毒载体的构建及鉴定[J].昆明医学院学报,2011,32(3):16-20. |
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| 作者姓名： | 尹倩 孙桂芹 庄艳艳 钟梅 苏桂栋 |
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| 作者单位： | 南方医科大学南方医院妇产科,广东,广州,510515 |
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| 基金项目： | 广东省自然科学基金资助项目 |
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| 摘 要： | 目的以人乳头瘤病毒（human papillomavirus,HPV）16型E6基因为靶点,构建小发夹状RNA（small hairpin RNA,shRNA）慢病毒载体.方法将HPV 16E6基因的特异性序列,应用基因重组技术插入到包含U6启动子及绿色荧光蛋白（GFP）报告基因的慢病毒载体pGCL-GFP中.重组质粒命名为pGCL-GFP-HPV16-E,测序鉴定后与慢病毒包装质粒pHelper 1.0及pHelper 2.0通过lipofectamine 2000共转染293T细胞,病毒液根据绿色荧光蛋白（GFP）表达水平测定滴度.结果 PCR扩增和测序结果证实HPV16 E6 shRNA核苷酸链序列插入正确,包装慢病毒产生病毒悬液,测定滴度为2×109 TU/mL.结论应用基因重组技术等方法,可成功构建HPV16 E6 shRNA慢病毒载体.
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| 关 键 词： | 人乳头瘤病毒 小发夹状RNA E6 慢病毒 |
Construction and Identification of Short Hairpin RNA Lentiviral Vector of Human Papillomavirus (HPV) 16 E6 |
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| YIN Qian,SUN Gui-qin,ZHUANG Yan-yan,ZHONG Mei,SU Gui-dong.Construction and Identification of Short Hairpin RNA Lentiviral Vector of Human Papillomavirus (HPV) 16 E6[J].Journal of Kunming Medical College,2011,32(3):16-20. |
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| Authors: | YIN Qian SUN Gui-qin ZHUANG Yan-yan ZHONG Mei SU Gui-dong |
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| Institution: | YIN Qian,SUN Gui-qin,ZHUANG Yan-yan,ZHONG Mei,SU Gui-dong(Dept.of Obstetrics and Gynecology,Nanfang Hospital,Southern Medical University,Guangzhou Guangdong 510515,China) |
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| Abstract: | Objective To establish a lentiviral vector-mediated RNA interference(RNAi)of Human Papillomavirus(HPV)16 E6 gene.Methods Using genetic recombination technology,a pair of complementary small hairpin RNA(shRNA)oligonucleotides targeting the HPV16 E6 gene was cloned into lentiviral expression vector pGCL-GFP,which contained U6 promoter and green fluorescent protein reporter.The recombined plasmid named pGCL-GFP-vshHPV 16 E6 was confirmed by polymerase chain reaction and sequencing,and was cotransfected with pH... |
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| Keywords: | E6 |
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