Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (27): 5071-5074.doi: 10.3969/j.issn.1673-8225.2010.27.030

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Construction of human sTRAIL vector and its expression in dermis-derived mesenchymal stem cells 

Miao Dian-nan1, Liang Yu-jia1, Yan Guo-he2, Dong Shi-wu3, Dai Xiao-tian1, Xiong Wei1   

  1. 1 Department of Respiratory Diseases, Southwest Hospital, Third Military Medical University, Chongqing  400038, China; 2 State Key Laboratory of Trauma, Burns and Combined Injury, Department of Nuclear Prevention Medicine, College of Military Preventive Medicine, 3 Department of Anatomy, State Key Laboratory for Biomechanics & Tissue Engineering, Ministry of Education, Third Military Medical University, Chongqing  400038, China
  • Online:2010-07-02 Published:2010-07-02
  • Contact: Xiong Wei, Doctor, Associate professor, Department of Respiratory Diseases, Southwest Hospital, Third Military Medical University, Chongqing 400038, China xiongwei64@126.com
  • About author:Mao Dian-nan★, Studying for master’s degree, Attending physician, Department of Respiratory Diseases, Southwest Hospital, Third Military Medical University, Chongqing 400038, China mdn7310@163.com
  • Supported by:

    the National Natural Science Foundation of China, No. 30770929*

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Abstract:

BACKGROUND: Based on the antineoplastic features of tumour necrosis factor (TNF) soluble related apoptosis inducing ligand (sTRAIL), sTRAIL gene was cloned and its eukaryotic expression vector was constructed, which can provide a foundation for apoptotic study of tumour cells.
OBJECTIVE: To construct pEGFP-N1/sTRAIL eukaryotic expression vector and express it into dermis-derived mesenchymal stem cells (dMSCs).
METHODS: cDNA fragment encoding human sTRAIL gene were amplified by reverse transcriptase-polymerase chain reaction (RT-PCR) with the total mRNA isolated from human placenta tissue as template. The PCR amplified fragment of sTRAIL gene was first cloned into Pmd18-T vector, and then subcloned into the recombinant eukaryotic expression vector pEGFP-N1 to construct pEGFP-N1/sTRAIL plasmid after sequencing. Subsequently, plasmid DNA of pEGFP-N1/sTRAIL was transfected into dMSCs with the help of Fugene 6 transfection reagent. The growth of dMSCs and expression of transfected sTRAIL in dMSCs were observed under an inverted microscope. The dMSCs were identified by RT-PCR.
RESULTS AND CONCLUSION: cDNA sequence encoding human sTRAIL gene was successfully cloned, and recombinant eukaryotic expression vector pEGFP-N1/sTRAIL was constructed. The expression of sTRAIL in dMSCs was obtained with the transfecton of pEGFP-N1/sTRAIL plasmid. Under an inverted microscope, the transfected DMSCs could be seen grown well. cDNA sequence encoding human sTRAIL gene was successfully cloned into pEGFP-N1. The transient expression of sTRAIL gene in dMSCs has been realized.

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