首页|FLIP腺病毒表达载体的构建及其在大鼠肺血管内皮细胞中的表达

FLIP腺病毒表达载体的构建及其在大鼠肺血管内皮细胞中的表达

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目的:构建FLIP(FLICE-inhibitory protein)腺病毒表达载体并观察该载体在大鼠肺血管内皮细胞(PVEC)中的表达情况,为研究FLIP蛋白的抗凋亡作用打下基础. 方法:利用含有大鼠FLIP基因的原始载体,通过Admax腺病毒包装系统,构建表达大鼠FLIP基因的重组腺病毒Ad-FLIP;原代培养并鉴定大鼠PVEC.利用成功构建的重组腺病毒Ad-FLIP感染大鼠PVEC.通过逆转录酶一多聚酶链反应(reverse tran-seriptase-polymerase chain reaction,RT-PeR)和Western Blot分别检测经重组腺病毒Ad-FLIP感染后大鼠PVEC及对照组细胞中FLIP基凶mRNA和蛋白表达水平. 结果:成功构建含有大鼠FLIP基因的重组腺病毒Ad-FIJP;经Ⅷ因子间接免疫荧光鉴定原代培养大鼠PVEC纯度达90%以上;大鼠PVEC经重组腺病毒Ad-FLIP感染后24 h即检测到FLIP mRNA和蛋白表达水平明显增高. 结论:重组腺病毒Ad-FLIP可有效提高大鼠PVEC中FLIP基因的表达水平.
Construction of adenoviral expression vector carrying FLIP gene and its expression in rat pulmonary vascular endothelial cells
AIM: To construct an adenoviral vector carrying rat FLIP (FLICE-inhibitory protein) gene and to observe its expression in rat pulmonary vascular endothelial cells (PVEC). METHODS: Recombined adenovirus Ad-FLIP was constructed by Ad-max system. The PVECs of rats were isolated, primarily cultured in vitro and infected with Ad-FLIP. The expression of FLIP was detected by Western blot and RT-PCR. RESULTS: FLIP gene was successfully cloned and incorporated into recombinant adenovirus Ad-FLIP. The expression of FLIP gene in the infected cells was detected by RT-PCR. The cells were infected with Ad-FLIP and FLIP protein was detected in the ingredients ofcell disruption. CONCLUSION: The recombinant adenoviral vector carrying rat FLIP gene is successfully constructed and Ad-FLIP efficiently elevates the expression of FLIP gene and protein in cultured rat PVECs.

FLIPadenoviralvascular endothelial cellsALL/ARDS

屈朔瑶、欧阳海峰、遆新宇、薛彦、吴昌归

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第四军医大学西京医院呼吸内科,陕西,西安,710033

FLIP 腺病毒 肺血管内皮细胞 ALI/ARDS

2009

第四军医大学学报
第四军医大学

第四军医大学学报

CSTPCDCSCD北大核心
影响因子:0.599
ISSN:1000-2790
年,卷(期):2009.30(18)
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