| 张英杰,陈剑,孙彦春.反义重组腺病毒质粒pAdxsi-GFP-hVEGF的构建与鉴定[J].济宁医学院学报,2011,(2):81-84,87 |
| 反义重组腺病毒质粒pAdxsi-GFP-hVEGF的构建与鉴定 |
| Construction and identification of the Adenoviral vector pAdxsi-GFP-hVEGF |
| 投稿时间：2011-01-05 |
| DOI：10.3969/j.issn.1000-9760.2011.02.002 |
| 中文关键词: 腺病毒|反义核酸|血管内皮生长因子|质粒 |
| 英文关键词: adenovirus|antisense|VEGF|plasmids |
| 基金项目: |
| 作者 | 单位 | | 张英杰 | 济宁医学院附属第一人民医院, 山东济宁 272011 | | 陈剑 | 济宁医学院附属第一人民医院, 山东济宁 272011 | | 孙彦春 | 济宁医学院附属第一人民医院, 山东济宁 272011 |
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| 中文摘要: |
| 目的 构建反义重组腺病毒质粒pAdxsi-GFP-hVEGF。方法 从人外周血细胞中提取总RNA,RT-PCR法反转录合成人cDNA文库。设计带有酶切位点的引物,调取VEGF目的基因,插入pShuttle-GFP-CMV穿梭载体。经鉴定序列正确后,将VEGF目的基因构建至亚克隆重组腺病毒质粒pAdxsi中。结果 成功构建反义重组腺病毒质粒pAdxsi-GFP-hVEGF,经酶切鉴定,目的基因分子量与预计的相同,测序插入载体部位正确。结论 反义重组腺病毒质粒pAdxsi-GFP-hVEGF构建正确、稳定,为进一步抑制人VEGF基因的表达和体内外实验打下了良好的基础。 |
| 英文摘要: |
| Objective To construct the Adenoviral vector pAdxsi-GFP-hVEGF.Methods Total RNA was isolated from blood cells and cDNA was constructed by RT-PCR, a pair of primers were synthesized with restriction site to get human VEGF cDNA fragments, and VEGF fragments were inserted into the shuttle vector pShuttle-GFP-CMV.After the sequence was confirmed, the human VEGF fragments were subcloned into Adenoviral vector pAdxsi-GFP-hVEGF.Results Adenoviral vector pAdxsi-GFP-hVEGF was successfully constructed, and the molecular weight of the target gene equaled the excepted, and sequencing results indicated that Adenoviral vector pAdxsi-GFP-hVEGF contained correct recombinant.Conclusion Adenoviral vector pAdxsi-GFP-hVEGF is correct and stable, and it will be useful for further research of VEGF inhibition both in vivo and in vitro. |
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