利用Tet-on调控系统建立人肝癌HepG2Tet-on细胞系
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金炜东

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Establishment of HepG2Tet-on cell line controlled by the Tet-on regulatory system
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    摘要:

    目的:构建可以用强力霉素调控表达的人肝癌HepG2Tet-on细胞系,为进一步研究肝癌相关基因功能奠定基础。
    方法:用脂质体转染法将pWHE146质粒转染到人肝癌HepG2细胞中,用G418筛选出稳定表达细胞克隆; 单克隆分别扩增后,瞬时转染pTRE-hyg-luc质粒; 强力霉素诱导表达后,检测荧光素酶表达活性,挑选出受强力霉素调控的低背景、高表达的HepG2Tet-on细胞株。
    结果:成功构建了一株受强力霉素调控的高表达低背景的HepG2Tet-on细胞株(诱导倍数达154.106倍)。
    结论:HepG2Tet-on细胞株可用于外源基因的真核调控高表达,为研究真核基因功能提供一种可靠的细胞株。

    Abstract:

    Objective:To establish tetracycline-controlled inducible system (Tet-on) in HepG2 cell for further research of the function of related gens.
    Methods :The HepG2 cells were transfected with pWHE146 vector by using liposome transfection reagent. The transfected cells were selected in medium containing G418, and G418-resistant clones were isolated. All individual G418-resistant clones were screened by transient transfection with plasmid pTRE-hyg-luc for clones with low background and high induction of luciferase in response to Dox.
    Results:One HepG2 cell line, which exhibited high levels of induction(154,106 times)and high gene expression levels, was obtained.
    Conclusions:The HepG2 cell line can be used to highly express eukaryotic gene and this Tet-on system is available for use in eukaryotic gene function studies.

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金炜东, 陈孝平, 陈钢.利用Tet-on调控系统建立人肝癌HepG2Tet-on细胞系[J].中国普通外科杂志,2009,18(2):10-.
DOI:10.7659/j. issn.1005-6947.2009.02.010

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  • 在线发布日期: 2009-02-25