| 野生型MLK3真核表达载体的构建及其表达 |
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| 引用本文： | 金磊,岳军,杜彩萍,侯筱宇.野生型MLK3真核表达载体的构建及其表达[J].徐州医学院学报,2010,30(2):82-85. |
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| 作者姓名： | 金磊 岳军 杜彩萍 侯筱宇 |
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| 作者单位： | 江苏省脑病生物信息重点实验室,徐州医学院生物化学与分子生物学研究中心,江苏,徐州,221002 |
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| 基金项目： | 教育部新世纪优秀人才支持计划，江苏省科技厅青年科技创新人才项目 |
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| 摘 要： | 目的构建野生型MLK3真核表达质粒并转染至COS-7细胞中表达。方法以大鼠海马总RNA为模板,通过RT-PCR分段扩增MLK3的序列,采用酶切、链接的方法先将目的基因克隆至pGEM-T,筛选正确的质粒后,再亚克隆至pcDNA3.1（＋）。以脂质体转染法将构建好的重组质粒转染至COS-7细胞,通过免疫印迹鉴定重组质粒的表达。结果限制性内切酶酶切和测序结果表明,扩增出的野生型MLK3的序列正确。免疫印迹显示,转染的重组质粒在相对分子质量92×103处有相应条带出现。结论成功构建了MLK3的真核表达质粒,重组质粒在COS-7细胞中高效表达。
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| 关 键 词： | MLK3 克隆 真核表达 |
Construction and expression of the wild-type MLK3 eukaryotic expression plasmid vector |
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| JIN Lei,YUE Jun,DU Caiping,HOU Xiaoyu.Construction and expression of the wild-type MLK3 eukaryotic expression plasmid vector[J].Acta Academiae Medicinae Xuzhou,2010,30(2):82-85. |
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| Authors: | JIN Lei YUE Jun DU Caiping HOU Xiaoyu |
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| Institution: | JIN Lei,YUE Jun,DU Caiping,HOU Xiaoyu (Jiangsu Key Laboratory of Brain Disease Bioinformation,Research Center for Biochemistry , Molecular Biology,Xuzhou Medical College,Xuzhou,Jiangsu 221002,China) |
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| Abstract: | Objective To construct the recombinant of wild-type MLK3 eukaryotic expression plasmid and to express the recombinant in transfected COS-7.Methods According to the cDNA sequence of MLK3(GenBank,BC081952),the sequence of wild-type MLK3 fragments were expanded by RT-PCR,using rat hippocampal total RNA as the template.Then these fragments were cloned into the plasmid of pcDNA3.1(+).The recombinant of pcDNA3.1-MLK3 was transfected into COS-7 cells by the mediation of lipofectamine reagents.Immunoblot method was... |
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| Keywords: | MLK3 |
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