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《Journal of Third Military Medical University》 2012-01
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Construction of three recombinant lentiviruses containing apoptin driven by survivin promoter of different lengths and comparison of their tumor suppressing efficency in vitro

Ye Feng1,Zhong Bo1,2,Dan Guorong1,Jiang Fan1,Sai Yan1,Zhao Jiqing1,Sun Huiqin3,Liang Houjie2,Zou Zhongmin1(1Institute of Toxicology,3State Key Laboratory of Trauma,Burns and Combined Injury,Institute of Combined Injury,College of Preventive Military Medicine,2Department of Oncology,Southwest Hospital,Third Military Medical University,Chongqing,400038,China)  
Objective To construct the recombinant lentivirus vectors using 3 survivin gene promoters(pSur) with a different length expressing 6 his-tagged apoptin cDNA and observe their tumor-suppressing effect after they were packed and human rectal cancer cells(SW480) were infected with them.Methods Three survivin gene promoters at a length of 160,270 and 987 bp expressing 6 his-tagged apoptin cDNA were cloned and linked to the pMD18-T vector on which survivin gene promoters and apoptin gene were assembled.The assembled expression box was constructed into the lentivirus vector FG12 modified with polyA.Recombinant lentiviruses were packed using the 3-plasmid packaging system.Tumor-suppressing effect of recombinant lentivirus vectors on SW480 cancer cells was observed while apoptosis and cell cycle of SW480 cancer cells were detected by Annexin V-PE/7-AAD and flow cytometry on days 5 and 30 after SW480 cancer cells were infected with recombinant lentiviruses at an optimal MOI.Results The 3 recombinant lentivirus vectors were successfully constructed,and high titer lentiviruses were obtained after they were packed and concentrated.The infection rate of target cells for the purified lentivirus was over 80%.In vitro examination showed that the SW480 cancer cells expressed the apoptin protein with apoptosis or necrosis occurred on day 30 after they were infected with recombinant lentivirus.The tumor suppressing effect of recombinant lentivirus containing the 270 bp survivin promoter was more significant.Conclusion Recombinant lentivirus constructed with the 270 bp survivin promoter driving apoptin expression in target cells shows a rather good tumor-suppressing effect on human rectal cancer.
【Fund】: 重庆市自然科学基金(CSTC2007BA5011)~~
【CateGory Index】: R735.3
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