IRF3 shRNA腺病毒包装及其对RAW264.7细胞IRF3表达的抑制效应

被引：1

作者：

涂文娟 ^{[1
,2
]}

朱彤 ^{[1
]}

谈志丽 ^{[1
]}

刘亮明 ^{[1
]}

机构：

[1] 南京医科大学附属上海松江中心医院感染科/上海交通大学附属第一人民医院松江分院感染科

[2] 上海市同济医院消化内科

来源：

免疫学杂志 | 2016年 / 32卷 / 10期

关键词：

干扰素调节因子3; 短发夹RNA; 腺病毒; 病毒包装; RAW264.7细胞;

D O I：

10.13431/j.cnki.immunol.j.20160167

中图分类号：

R373 [人体病毒学（致病病毒）];

学科分类号：

摘要：

目的重组并包装干扰素调节因子3(interferon regulator factor 3,IRF3)短发夹RNA(short hairpin RNA,sh RNA)腺病毒,研究该病毒对RAW264.7细胞IRF3基因表达及对细胞增殖活性和吞噬功能的影响。方法采用酶切和连接方法构建p Adeno-U6-CMV-EGFP-IRF3 sh RNA质粒;借助Ad MaxTM病毒包装系统完成IRF3基因干扰腺病毒的重组与包装;IRF3基因表达分别采用real-time PCR和Western blot分析方法检测;RAW264.7细胞的增殖活性和吞噬功能分别采用CCK8分析和吞墨试验检测。结果 IRF3基因的干扰片段及对照序列被正确插入p Adeno-U6-CMV-EGFP质粒AgeⅠ和Eco RⅠ酶切位点之间,经PCR扩增及DNA测序证实;RAW264.7细胞组成性表达IRF3基因,针对IRF3基因后部序列的干扰腺病毒Y2147明显抑制了IRF3 m RNA和蛋白质的表达,而针对IRF3基因前部及中部序列及对照序的干扰腺病毒对IRF3的表达无明显沉默或封闭效应;腺病毒Y2147对RAW264.7细胞的增殖活性及吞噬功能无明显不良影响。结论成功构建并包装了IRF3 sh RNA腺病毒,该病毒能有效抑制RAW264.7细胞IRF3 m RNA和蛋白表达,且对细胞的增殖活性和吞噬能力无不良影响。

引用

页码：857 / 863

页数：7

共 19 条

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