中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (41): 7693-7696.doi: 10.3969/j.issn.1673-8225.2011.41.023

• 组织构建细胞学实验 cytology experiments in tissue construction • 上一篇    下一篇

人ski基因真核表达载体的构建及促细胞增殖作用

彭  艳，李  平，刘  苹，陈  磊，赵晓光，周元国   

  1. 解放军第三军医大学附属大坪医院野战外科研究所分子生物学中心，创伤、烧伤与复合伤国家重点实验室，重庆市  400042
  • 收稿日期:2011-04-01 修回日期:2011-06-12 出版日期:2011-10-08 发布日期:2011-10-08
  • 通讯作者: 周元国，研究员，解放军第三军医大学附属大坪医院野战外科研究所分子生物学中心，创伤、烧伤与复合伤国家重点实验室，重庆市 400042 ygzhou@cta.cq. cn
  • 作者简介:彭艳★，女，1982年生，安徽省宿州市人，汉族，2009年徐州医学院毕业，硕士，主要从事创伤愈合方面的研究。 pengyan1006@ 126.com
  • 基金资助:

    创伤、烧伤与复合伤国家重点实验室自主课题(No. SKLZZ200805)，课题名称：具有促进创伤愈合和抑制瘢痕增生双重作用的人ski基因的应用研究。

Construction of eukaryotic expression vecter for human ski gene and observation of its role in promoting cell proliferation

Peng Yan, Li Ping, Liu Ping, Chen Lei, Zhao Xiao-guang, Zhou Yuan-guo   

  1. State Key Laboratory of Trauma, Burns and Combined Injury, Center for Molecular Biology, Research Institute of Surgery and Daping Hospital, Third Military Medical University of Chinese PLA, Chongqing   400042, China
  • Received:2011-04-01 Revised:2011-06-12 Online:2011-10-08 Published:2011-10-08
  • Contact: Zhou Yuan-guo, Researcher, State Key Laboratory of Trauma, Burns and Combined Injury, Center for Molecular Biology, Research Institute of Surgery and Daping Hospital,Third Military Medical University of Chinese PLA, Chongqing 400042, China ygzhou@cta.cq.cn
  • About author:Peng Yan★, Master, State Key Laboratory of Trauma, Burns and Combined Injury, Center for Molecular Biology, Research Institute of Surgery And Daping Hospital,Third Military Medical University of Chinese PLA, Chongqing 400042, China pengyan1006@126. com
  • Supported by:

    An Independent Topic of State Key Laboratory of Trauma, Burns and Combined Injury, No. SKLZZ200805*

PDF

517

被引次数

3

摘要：

背景：c-Ski既可促进组织修复，又能降低瘢痕形成，有望成为一个全新的基因治疗药物。
目的：构建人ski基因的真核表达载体，并对其促成纤维细胞增殖效果进行验证。
方法：RT-PCR法从人包皮培养原代成纤维细胞总RNA中扩增出人ski基因，连同真核表达启动子CMV克隆到pUC118载体上，酶切和测序验证后，应用脂质体将其转染到培养的大鼠皮肤成纤维细胞中。
结果与结论：酶切和测序结果证实表达质粒构建成功，Western blot显示Ski蛋白在转染细胞中成功表达，且可显著提高转染细胞的增殖效应，说明以pUC118为骨架的重组ski表达质粒具有促进大鼠皮肤成纤维细胞增殖的作用。

关键词: 载体构建, ski, pUC118, 成纤维细胞, 增殖

Abstract:

BACKGROUND: c-Ski can promote tissue repair and alleviate scar formation, which is expected to be a new gene therapeutic drugs.
OBJECTIVE: To construct eukaryotic expression plasmid of human ski gene and to investigate its proliferative role in fibroblasts.
METHODS: The ski gene was amplified by polymerase chain reaction (PCR) from human foreskin cultured fibroblast and was sub-cloned into pUC118 with CMV eukaryotic promoter. The recombinant pUC118-Ski plasmids were transfected into rat skin fibroblasts using lipofectamine TM 2000 and then its protein expression and effect of the proliferation were detected.
RESULTS AND CONCLUSION: pUC118-Ski vector was constructed successfully which was confirmed by restriction endonuclease digestion and DNA sequencing. It expressed successfully and significantly improved the effect of the proliferation in transfected fibroblasts. Recombinant eukaryotic expression plasmid pUC118-Ski can improve the proliferation of rat skin fibroblasts.

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