中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (10): 1755-1758.doi: 10.3969/j.issn.1673-8225.2011.10.010

• 干细胞转基因表达 transgenic expression in stem cells • 上一篇    下一篇

人胚胎干细胞外源基因的瞬时表达效率

路  君，陈宗峰，黄粱浒，谭建明   

  1. 解放军南京军区福州总医院福建省移植生物学重点实验室，福建省福州市  350025
  • 收稿日期:2010-11-28 修回日期:2011-01-28 出版日期:2011-03-05 发布日期:2011-03-05
  • 通讯作者: 谭建明，博士生导师，教授，解放军南京军区福州总医院福建省移植生物学重点实验室，福建省福州市350025 doctortjm@yahoo.com.cn
  • 作者简介:路君☆，女，1981年生，福建省福州市人，汉族，2008年复旦大学毕业，博士，助理研究员，主要从事干细胞分化方面的研究。 junlu.heather@yahoo.com
  • 基金资助:

    福建省自然科学基金项目(2010J05078，2010J01217)。

Transient expression efficiency of foreign gene in human embryonic stem cells

Lu Jun, Chen Zong-feng, Huang Liang-hu, Tan Jian-ming   

  1. Fujian Provincial Key Laboratory of Transplant Biology, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA, Fuzhou  350025, Fujian Province, China
  • Received:2010-11-28 Revised:2011-01-28 Online:2011-03-05 Published:2011-03-05
  • Contact: Tan Jian-ming, Doctoral supervisor, Professor, Fujian Provincial Key Laboratory of Transplant Biology, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA, Fuzhou 350025, Fujian Province, China doctortjm@yahoo.com.cn
  • About author:Lu Jun☆, Doctor, Assistant researcher, Fujian Provincial Key Laboratory of Transplant Biology, Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA, Fuzhou 350025, Fujian Province, China junlu.heather@yahoo.com
  • Supported by:

    Natural Science Foundation of Fujian Province, No. 2010J05078, 2010J01217*

PDF

416

被引次数

2

摘要：

背景：人类胚胎干细胞的修饰和操作技术多种多样，它们在效率、可靠性及安全性方面各有不同。
目的：对比观察两种化学转染试剂转染含不同启动子的增强型绿色荧光蛋白表达载体在人胚胎干细胞中的表达效率。
方法：采用Fugene HD和lipofectamine2000分别转染无滋养层培养的H9细胞，荧光显微镜下观察阳性克隆，流式细胞术分析不同载体pCAG-EGFP、pEGFP-N3在H9细胞中的表达效率。
结果与结论：Fugene HD转染的pCAG-EGFP、pEGFP-N3两种载体在H9细胞中的表达效率分别为(42.45±3.32)%、 (25.95±1.91)%，差异有显著性意义(P < 0.05)。lipofectamine2000转染的pCAG-EGFP、pEGFP-N3两种载体在H9细胞中的表达效率分别为(1.94±0.18)%、(1.49±0.33)%。采用Fugene HD转染的表达效率均高于lipofectamine2000转染的表达效率(P < 0.05)。结果显示在人胚胎干细胞的H9细胞系中，Fugene HD的转染效率显著高于lipofectamine2000；CAG启动子驱动的增强型绿色荧光蛋白表达效率显著高于CMV启动子。

关键词: 人胚胎干细胞, 基因转染, 表达效率, 启动子, 增强型绿色荧光蛋白

Abstract:

BACKGROUND: Technologies designed to allow modification and manipulation of human embryonic stem cells (hESCs) is numerous and various in the complexity of their efficiency, reliability and safety.
OBJECTIVE: To compare effect of different enhanced green fluorescent protein (EGFP) expression vectors with different promoter on expression efficiency in human embryonic stem cells (hESCs).
METHODS: The H9 cells cultured with non trophoblast were transfected with Fugene HD and lipofectamine 2000, positive colonies were detected under fluorescence microscope, and the expression efficiency were measured using flow cytometry analysis between different transfect vectors such as pCAG-EGFP and pEGFP-N3.
RESULTS AND CONCLUSION: Expression efficiencies of pCAG-EGFP and pEGFP-N3 were (42.45±3.32)% and (25.95±1.91)% respectively in Fugene HD transfected hESCs, and they had statistically difference (P < 0.05). It was (1.94±0.18)% and (1.49±0.33)% in lipofectamine 2000. The expression efficiencies of pCAG-EGFP and pEGFP-N3 transfected with Fugene HD were higher than that transfected with lipofectamine 2000 (P < 0.05).These results indicate that Fugene HD have the greater expression efficiency compared to lipofectamine 2000 in hESC line H9; expression efficiency of EGFP driven by CAG promoter is higher than that by CMV promoter.

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