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中华普通外科学文献(电子版) ›› 2008, Vol. 02 ›› Issue (05) : 349 -352. doi: 10.3877/cma.j.issn.1674-0793.2008.05.005

论著

单核细胞趋化蛋白-1小分子干扰RNA真核表达质粒的构建及意义
廖艺1, 胡作军1,(), 吕伟明1, 王嘉励2, 胡玲玉1, 林勇杰1, 王深明1   
  1. 1.510080 广州,中山大学附属第一医院血管外科
    2.中山大学医学院生化教研室
  • 收稿日期:2008-03-07 出版日期:2008-10-01
  • 通信作者: 胡作军
  • 基金资助:
    广东省科技计划重点引导项目资助(2005B31201001)教育部高等学校博士点专项基金项目资助(20050558053)

Construction eukaryotic expression vector of MCP-1 siRNA

Yi LIAO1, Zuo-jun HU1,(), Wei-ming LV1, Jia-li WANG1, Ling-yu HU1, Yong-jie LIN1, Shen-ming WANG1   

  1. 1.Department of vascular surgery, The First Affiliated Hospital of Sun Yat-sen University,Guangzhou 510080,China
  • Received:2008-03-07 Published:2008-10-01
  • Corresponding author: Zuo-jun HU
引用本文:

廖艺, 胡作军, 吕伟明, 王嘉励, 胡玲玉, 林勇杰, 王深明. 单核细胞趋化蛋白-1小分子干扰RNA真核表达质粒的构建及意义[J/OL]. 中华普通外科学文献(电子版), 2008, 02(05): 349-352.

Yi LIAO, Zuo-jun HU, Wei-ming LV, Jia-li WANG, Ling-yu HU, Yong-jie LIN, Shen-ming WANG. Construction eukaryotic expression vector of MCP-1 siRNA[J/OL]. Chinese Archives of General Surgery(Electronic Edition), 2008, 02(05): 349-352.

目的

构建单核细胞趋化蛋白-1(MCP-1)小分子干扰RNA(siRNA)表达载体,为进一步研究其对动脉粥样硬化(AS)的防治作用提供手段。

方法

设计并合成两端含有酶切位点两条DNA序列,经退火成互补双链,再克隆至载体pSilencer2.0-U6中构建重组表达载体,转化DH-5α菌株,提取质粒行双酶切鉴定,并进行序列测定。

结果

双酶切证实MCP-1 siRNA表达载体克隆构建成功,插入片段测序结果与合成的siRNA结果一致。

结论

成功构建MCP-siRNA表达载体,为动脉粥样硬化的防治奠定实验基础。

Objective

To construct the small interfering RNA (siRNA) expression vector for monocyte chemotactite protein-1(MCP-1)and provide ways for artery stenosis treatment.

Methods

Two DNA sequences containing the sites of restriction endonuclease at both ends were designed and synthesized.The eomplement form was obtained by annealing and being cloned into vector pSilencer2.0-U6 and the recombinant plasmid was transformed into strain DH-5α. The plasmid identified by restriction enzyme was used for sequencing.

Results

MCP-1 siRNA expression vector was successfully constructed and identified by double endonuclease digestion.Sequence analysis of the inserted fragment revealed the salne sequence as that of the synthesized siRNA oligonucleotides.

Conclusion

MCP-1 siRNA expression vector has been successfully constructed,which lays the basis for its application in the prevention and treatment of artery stenosis.

图1 psilencer2.0-U6-siMCP-1 重组质粒的构建图
图2 MCP-1:sh1
图3 MCP-1:sh2
图4 阳性重组体(psilencer2.0-U6-siMCP-1)序列测定结果(sh1)
图5 阳性重组体(psilencer2.0-U6-siMCP-1)序列测定结果(sh2)
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