| VEGF121和BMP2双基因共表达重组腺病毒载体的构建及其在HEK293中的表达 |
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| 引用本文： | 栗刚,吴秀成,钟声,王巍,李媛,刘丹平.VEGF121和BMP2双基因共表达重组腺病毒载体的构建及其在HEK293中的表达[J].山东医药,2011,51(10):4-6. |
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| 作者姓名： | 栗刚 吴秀成 钟声 王巍 李媛 刘丹平 |
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| 作者单位： | 1. 辽宁医学院附属第一医院,辽宁锦州,121000
2. 辽源市中心医院 |
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| 基金项目： | 国家自然科学基金资助项目 |
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| 摘 要： | 目的构建人血管内皮生长因子121（VEGF121）与人骨形态发生蛋白2（BMP2）双基因共表达腺病毒载体Adv-BMP2-IRES-VEGF121,并观察其在人胚肾细胞株（HEK293）中的表达情况。方法对腺病毒质粒pShuttle-CMV-BMP2的目的基因BMP2进行PCR扩增。腺病毒质粒pShuttle-CMV-VEGF121-IRES-hrGFP-1经Kpn I/Xba I酶切后,将BMP2片段定向导入pShuttle-CMV-VEGF121-IRES,构建pShuttle-CMV-V EGF121-IRES-BMP2,并注入大肠杆菌DH5a中扩增,提取质粒。通过酶切分析、PCR检测和序列分析进行鉴定。将构建所得的质粒转染HEK293,采用RT-PCR法检测HEK293中的BMP2、VEGF121 mRNA,Western blot法检测其蛋白。结果成功构建了Adv-BMP2-IRES-VEGF121。酶切分析及DNA序列测定证实重组质粒构建正确。质粒转染后的HEK293 BMP2和VEGF121表达阳性。结论成功构建了Adv-BMP2-IRES-VEGF121,其转染HEK293后,VEGF121、BMP2在HEK293中共表达阳性。
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| 关 键 词： | 人血管内皮生长因子121 人骨形态蛋白2 腺病毒载体 质粒 基因工程 |
Construction and identification of recombinant adenovirus vector co-expressing VEGF121 and BMP2 genes and its expression in HEK293 cells |
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| LI Gang,WU Xiu-cheng,ZHONG Sheng,WANG Wei,LI Yuan,LIU Dan-ping.Construction and identification of recombinant adenovirus vector co-expressing VEGF121 and BMP2 genes and its expression in HEK293 cells[J].Shandong Medical Journal,2011,51(10):4-6. |
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| Authors: | LI Gang WU Xiu-cheng ZHONG Sheng WANG Wei LI Yuan LIU Dan-ping |
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| Institution: | LI Gang1,WU Xiu-cheng,ZHONG Sheng,WANG Wei,LI Yuan,LIU Dan-ping(1 First Affiliated Hospital of Liaoning Medical University,Jinzhou 121001,P.R.China) |
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| Abstract: | Objective To construct and identify the adenovirus shuttle plasmid pShuttle-CMV-VEGF121-IRES-BMP2 and its express in HEK293 cells.Methods The DNA fragments of human BMP2 gene were changed restriction sites and subcloned by PCR.The human BMP2 genes and pShuttle-CMV-VEGF121-IRES were ligated into the plasmid by directional cloning method.The inserted target genes in the plasmid were verified by restriction enzyme digestion and nucleotide sequencing.The correct recombinant express plasmid was transfected to HE... |
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| Keywords: | human vascular endothelial growth factor 121 human bone morphogenetic protein 2 adenvirus vector plasimid genetic engineering |
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