| pcDNA3/BDNF真核表达载体的构建 |
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| 引用本文： | 孟步亮,徐丹,刘佳,李力燕,王廷华. pcDNA3/BDNF真核表达载体的构建[J]. 中国组织工程研究与临床康复, 2011, 15(2): 241-244. DOI: 10.3969/j.issn.1673-8225.2011.02.012 |
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| 作者姓名： | 孟步亮 徐丹 刘佳 李力燕 王廷华 |
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| 作者单位： | 1. 昆明医学院解剖学教研室,云南省昆明市,650031
2. 昆明医学院神经科学研究所,云南省昆明市,650031 |
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| 摘 要： | 背景:脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)作用广泛,但属于生物大分子,不能通过血脑屏障。基因治疗是目前解决脑源性神经营养因子给药途径最有希望的方案。目的:拟构建大鼠脑源性神经营养因子基因真核表达载体。方法:采用反转录聚合酶链式反应技术从SD大鼠脑组织提取总RNA,扩增脑源性神经营养因子基因cDNA序列,并将其克隆到真核表达载体pcDNA3中,分别取10g质粒pcDNA3和纯化的目的基因分别进行EcoRⅠ、xhoⅠ双酶切。将目的基因片段和pcDNA3载体连接,转入感受态DH5α细胞中,经酶切鉴定后送上海博亚生物技术有限公司测序。结果与结论:RT-PCR产物为749bp的特异片段,重组质粒pcDNA3/BDNF酶切后产生749bp和5446bp的片段,DNA测序证实749bp片段的碱基序列与大鼠脑源性神经营养因子基因序列完全一致,成功构建了pcDNA3/BDNF重组质粒。
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| 关 键 词： | 脑源性神经营养因子 真核表达载体 重组质粒 克隆 基因表达 |
Construction of pcDNA3/BDNF eukaryotic expression vectors |
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| Meng Bu-liang,Xu Dan,Liu Jia,Li Li-yan,Wang Ting-hua. Construction of pcDNA3/BDNF eukaryotic expression vectors[J]. Journal of Clinical Rehabilitative Tissue Engineering Research, 2011, 15(2): 241-244. DOI: 10.3969/j.issn.1673-8225.2011.02.012 |
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| Authors: | Meng Bu-liang Xu Dan Liu Jia Li Li-yan Wang Ting-hua |
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| Affiliation: | Meng Bu-liang1,Xu Dan2,Liu Jia2,Li Li-yan2,Wang Ting-hua21Department of Anatomy,2Institute of Neuroscience,Kunming Medical College,Kunming 650031,Yunnan Province,China |
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| Abstract: | BACKGROUND:Brain-derived neurotrophic factor (BDNF) is biomacromolecule,which can not pass through the blood-brain barrier.Now gene therapy is the most promising program to solve route of administrationOBJECTIVE:To construct the eukaryotic expression recombinant plasmid pcDNA3/BDNF.METHODS:Total RNA was extracted from Sprague Dawley rats using RT-PCR.By gene recombination technique,rat BDNF coding sequence was inserted into eukaryotic expression vector pcDNA3.The fragment of target gene was connected with p... |
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