Transfection of mouse bone marrow mesenchymal stem cells with liposome mediated cytosine deaminase genes Wu Chen-huan1, Wang Hong-fei1, Song Fei21Department of Orthopedics, 2Department of Neurosurgery, Second Affiliated Hospital of Dalian Medical University, Dalian 116023, Liaoning Province, ChinaBACKGROUND: The particular bystander effect of suicide gene can remarkably kill tumor cells. It can be used together with radiotherapy as well as immune gene therapy, and overcome the defect of low gene transduction efficiency. Cytosine deaminas(CD) can generate a powerful bystander effect. OBJECTIVE: To observe the effect of a eukaryotic expression plasmid pIRES2-AcGFP1-CD mediated by liposome transfectedinto bone marrow mesenchymal stem cells (BMSCs) and its gene expression. DESIGN, TIME AND SETTING: A cytologic experiment of genetic level was performed at Research and Development Center oStem Cell and Tissue Engineer, Dalian University of Technology from May to December 2007. MATERIALS: A total of six C57BL mice of SPF degree were provided by Experimental Animal Center of Dalian Medical University. E. DH5α was provided by Research and Development Center of Stem Cell and Tissue Engineer, Dalian University oTechnology. LipofectamineTM 2000 was the product of Invitrogen, China. METHODS: The DNA plasmids were extracted from transformed into the competent E. DH5α. pIRES2-AcGFP1-CD plasmid waidentified by BamHI/XhoI double digestion. The BMSCs from mouse femur and tibia were cultured and purified by adhesion method in vitro. Signal cell suspension prepared by BMSCs of the third generation was cultured by adding fluorescence-labeledCD44, CD45, CD90 and CD105 antibodies. BMSCs of the third generation were transfected by lipofectamine 2000 mediation. MAIN OUTCOME MEASURES: Identification of recombinant plasmids. The expressions of surface markers on BMSCs were detected by fluoroscopy. The expressions of CD gene were observed after transfection. RESULTS: After agarose gel electrophoresis, a band appeared at 1.0-1.5 kb of the digested products of pIRES2-AcGFP1-CD plasmids and accorded with the length of CD gene in the length. The cells were negative for CD45, and positive for CD44, CD90and CD105. Under the fluorescent inverted phase contrast microscope, the expressions of green fluorescent protein in the BMSCs were found at 36 hours after transfection. At 48 hours after transfection, the expressions of green fluorescent protein remained and the intensity increased obviously. CONCLUSION: The expression of CD gene mediated by liposome in BMSCs is successful, and it reaches the peak 48 hours after transfection. 【CateGory Index】： R329;R73-3 Download(CAJ format) Download(PDF format) CAJViewer7.0 supports all the CNKI file formats; AdobeReader only supports the PDF format. Ai-Qing Zheng, Tianjin Medical University, Tianjin 300070, China Ai-Qing Zheng, Xian-Rang Song, Ling Wei, Xing-Wu Wang, Cancer Research Center, Shandong Cancer Hospital, Jinan 250117, Shandong Province, China Jin-Ming Yu, Department of Radiation Oncology, Shandong Cancer Hospital, Jinan 250117, Shandong Province, China;Liposome transfected to plasmid-encoding endostatin gene combined with radiotherapy inhibits liver cancer growth in nude mice[J];世界胃肠病学杂志(英文版);2005-28 Song Fei1,2,Xing Qi3,Ji Guang-chun2,Ma Yu-fang4,Ma Xue-hu11College of Environmental Life Science,Dalian University of Technology,Dalian 116027,Liaoning Province,China;2Department of Neurosurgery,Second Clinical College,Dalian Medical University,Dalian 116023,Liaoning Province,China;3Department of Neurosurgery,First Clinical College,Dalian Medical University,Dalian 116011,Liaoning Province,China;4Department of Biochemistry and Molecular Biology Dalian Medical University,Dalian 116044,Liaoning Province,China;Transfection of mouse bone marrow mesenchymal stem cells with Lipofectamine-mediated cytosine deaminase genes[J];Journal of Clinical Rehabilitative Tissue Engineering Research;2009-49