Chinese Journal of Tissue Engineering Research ›› 2012, Vol. 16 ›› Issue (28): 5255-5259.doi: 10.3969/j.issn.2095-4344.2012.28.026

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Construction of a short hairpin RNA interference lentiviral vector of SIRT1 gene and detection of interference effect

Deng Zhao-ling1, Wang Yan2, Du Li-qing2, Liu Qiang2, Zhao Hui3, Chen Nai-yao1   

  1. 1Department of Hematology, Affiliated Hospital of Hebei United University, Tangshan 063000, Hebei Province, China;
    2Institute of Radiation Medicine, Chinese Academy of Medical Sciences, Tianjin Key Laboratory of Molecular Nuclear Medicine, Tianjin 300192, China;
    3Biochemistry and Food Technology School, Tianjin University of Commerce, Tianjin Key Laboratory of Food Science and Biotechnology, Tianjin 300134, China
  • Received:2011-12-01 Revised:2011-12-22 Online:2012-07-08 Published:2012-07-08
  • Contact: Chen Nai-yao, Chief physician, Professor, Master’s supervisor, Department of Hematology, Affiliated Hospital of Hebei United University, Tangshan 063000, Hebei Province, China nychenncmc@yahoo.com.cn
  • About author:Deng Zhao-ling★, Studying for master‘s degree, Department of Hematology, Affiliated Hospital of Hebei United University, Tangshan 063000, Hebei Province, China ling112711@163.com

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Abstract:

BACKGROUND: SIRT1 gene plays a very important role in cell energy metabolism, apoptosis and ageing, while other studies have found that SIRT1 may play a regulatory role in the inflammatory response.
OBJECTIVE: To construct short hairpin RNA interference lentiviral vector of SIRT1 gene and to evaluate their inhibitory effect in THP-1 cells.
METHODS: The SIRI1 target specific oligonucleotide sequence was designed, synthesized and connected into the pGCSIL-GFP vector digested by Age Ⅰ and EcoRⅠ 293T cells were packaged to produce the lentivirus, and then transfected with THP-1 cells. The inhibitory effect on SIRT1 gene was tested by the real-time quantitative PCR and Western Blot.
RESULTS AND CONCLUSION: The PCR and DNA sequencing of positive clones demonstrated that the lentivirus shRNA vector of SIRT1 gene was constructed successfully. The real-time quantitative PCR and Western Blot confirmed that the vector could suppress the expression level of SIRT1 at mRNA and protein levels.

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