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使用FLAG标签肽及慢病毒载体共同筛选小鼠foxp3基因RNA干扰的有效靶点
引用本文:史艳侠,韩文杰,彭柔君,张景航,黄慧强,姜文奇.使用FLAG标签肽及慢病毒载体共同筛选小鼠foxp3基因RNA干扰的有效靶点[J].中山大学学报(医学科学版),2007,28(6):641-644.
作者姓名:史艳侠  韩文杰  彭柔君  张景航  黄慧强  姜文奇
作者单位:华南肿瘤学国家重点实验室//中山大学肿瘤防治中心化疗科 广东广州510060(史艳侠,彭柔君,黄慧强,姜文奇),新乡医学院 河南新乡453003(韩文杰,张景航)
基金项目:国家自然科学基金;广东省卫生厅科研项目
摘    要: 【目的】筛选高效的foxp3 RNA干扰的靶点用于阻断CD4^+CD25^+ Treg细胞的免疫抑制功能。【方法】使用oligoengine软件设计foxp3基因shRNA序列,化学合成法合成4条shRNA序列,构建含foxp3的靶序列的慢病毒载体:构建出表达FLAG融合蛋白和目的基因的工具细胞293T细胞用于筛选RNA干扰的有效靶点。【结果】成功包装了4条foxp3基因靶序列相应的慢病毒颗粒;成功构建了表达融合蛋白和目的基因的293T工具细胞:在工具细胞上筛选出了2条RNAi效率分别为91.3%和95.6%的有效靶点。【结论】使用FLAG融合蛋白和慢病毒载体能够筛选出Foxp3基因的RNAi最佳靶点。

关 键 词:foxp3  FLAG蛋白  慢病毒载体  CD4^+CD25^+  Treg  RNA干扰
文章编号:1672-3554(2007)06-0641-05
收稿时间:2007-05-25;
修稿时间:2007年5月25日

Screening of RNA Intervening Targets of foxp3 Gene with FLAG Fusion Peptide and Lentivirus Vector
SHI Yan-xia,HAN Wen-jie,PENG Rou-jun,ZHANG Jing-hang,HUANG Hui-qiang,JIANG wen-qi.Screening of RNA Intervening Targets of foxp3 Gene with FLAG Fusion Peptide and Lentivirus Vector[J].Journal of Sun Yatsen University(Medical Sciences),2007,28(6):641-644.
Authors:SHI Yan-xia  HAN Wen-jie  PENG Rou-jun  ZHANG Jing-hang  HUANG Hui-qiang  JIANG wen-qi
Abstract:Objective] To screen the high effective RNA intervening targets of foxp3 gene to block the immunosuppression of CD4 CD25 Tregs. Method] Four shRNA sequences were designed by oligoengine software and synthesized by chemical method. Lentiviral vector that consists target sequences of foxp3 was constructed. The 293T tool cell line that express FLAG fusion protein and target gene was constructed to screen the effective targets for RNA interference. Result] Lentiviral vector that contains foxp3 cDNA sequences was constructed successfully. The 293T tool cells that express FLAG fusion protein and foxp3 gene fragments were constructed successfully. Two effective targets for RNAi were screened in 293T tool cells, the blocking efficiency were 91.3% and 95.6 % respectively. Conclusion] The optimal targets of Foxp3 gene for RNAi can be screened by FLAG fusion protein and lentivirus vector.
Keywords:foxp3
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