| 重组慢病毒载体介导HIV-1 Nef蛋白对原发性渗出性淋巴瘤细胞系和血管内皮细胞增殖作用影响的初探 |
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| 引用本文： | 朱小飞,秦 娣,周 峰,卫冰冰,卢 春.重组慢病毒载体介导HIV-1 Nef蛋白对原发性渗出性淋巴瘤细胞系和血管内皮细胞增殖作用影响的初探[J].南京医科大学学报,2011(2):131-137. |
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| 作者姓名： | 朱小飞 秦 娣 周 峰 卫冰冰 卢 春 |
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| 作者单位： | 南京医科大学微生物学与免疫学系; |
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| 基金项目： | 国家自然科学基金(30900064); 教育部高等学校博士点专项科研基金新教师基金(20093234120004); 江苏省高校自然科学基金(09KJB310007) |
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| 摘 要： | 目的:构建含有HIV-1负调控因子(Nef)基因的重组慢病毒表达载体,并检测Nef蛋白在人胚肾293T细胞、体腔渗出性淋巴瘤细胞BCBL-1和人血管内皮细胞EA.hy926中的表达情况及其对BCBL-1和EA.hy926细胞增殖的影响。方法:从本实验室已构建好的含Nef基因的重组真核表达质粒pCI-neo-Nef中扩增出Nef基因,插入到慢病毒载体pHAGE-CMV-MCS-Izs-Green中构建成pHAGE-Nef,利用脂质体将其与包装质粒psPAX2和包膜质粒pMD2.G共转染293T细胞。荧光显微镜下观察293T细胞中绿色荧光蛋白(GFP)的表达并收集培养上清,此上清经0.45μm滤器过滤后即获得慢病毒悬液。梯度稀释法测定慢病毒滴度。将重组慢病毒分别感染293T、BCBL-1和EA.hy926细胞,用Western blot检测Nef蛋白的表达。通过细胞增殖实验检测Nef蛋白对BCBL-1和EA.hy926细胞增殖的影响。结果:限制性内切酶鉴定和核酸序列测序证实成功构建了含HIV-1 Nef基因的重组慢病毒表达载体,病毒滴度为1×107 TU/ml。以重组慢病毒分别感染293T、BCBL-1和EA.h...
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| 关 键 词： | 慢病毒 Nef 卡波济肉瘤 卡波济肉瘤相关疱诊病毒 |
| 收稿时间： | 2010/9/15 0:00:00 |
Role of HIV-1 Nef protein carried by recombinant lentivirus in proliferation of primary effusion lymphoma cell line and vascular endothelial cells |
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| ZHU Xiao-fei,QIN Di,ZHOU Feng,WEI Bing-bing and LU Chun.Role of HIV-1 Nef protein carried by recombinant lentivirus in proliferation of primary effusion lymphoma cell line and vascular endothelial cells[J].Acta Universitatis Medicinalis Nanjing,2011(2):131-137. |
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| Authors: | ZHU Xiao-fei QIN Di ZHOU Feng WEI Bing-bing and LU Chun |
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| Institution: | ZHU Xiao-fei,QIN Di,ZHOU Feng,WEI Bing-bing,LU Chun*(Department of Microbiology and Immunology,NJMU,Nanjing 210029,China) |
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| Abstract: | Objective: To construct the recombinant lentivirus containing HIV-1 Nef gene and detect the protein expression in its target cell lines,such as 293T,BCBL-1 and EA.hy926,and explore the effect of Nef on cell proliferation.Methods:The fragment of Nef gene from expression vector pCI-neo-Nef was cloned into the lentivirus vector pHAGE-CMV-MCS-Izs-Green,then the recombinant vector pHAGE-Nef,packaging vector psPAX2 and envelope vector pMD2.G were cotransfected into the 293T cells.Culture media were harvested and ... |
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| Keywords: | recombinant lentivirus Nef KS KSHV |
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