人Slit2全长cDNA的真核表达及鉴定
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国家自然科学基金资助项目(30571873)


Eukaryotic expression and identification of human Slit2 gene in HEK-293 cell line
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    目的:对人神经轴突导向因子Slit2(hSlit2)全长cDNA进行真核表达并进行鉴定-方法:采用分段克隆的方法获得hSlit2全长cDNA的克隆,并构建hSlit2全长cDNA真核表达重组质粒pCMV-GFP-C/hSlit2,用磷酸钙共沉淀法将其转染到人胚肾细胞HEK-293细胞中,通过免疫印迹法鉴定其蛋白表达-结果:经酶切鉴定证实hSlit2全长cDNA的真核表达载体构建成功,免疫荧光蛋白和Western blot结果证实hSlit2全长cDNA在HEK-293细胞中的表达-结论:人神经轴突导向因子hSlit2真核表达载体构建成功,为进一步完善hSlit2蛋白及各水解片段功能研究提供了实验基础-

    Abstract:

    Objective:To construct the recombinant eukaryotic expression vector for human Slit2 full-length cDNA and express it in human embryonic kidney cells (HEK-293 cell line). Methods:The full length cDNA of hSlit2 was obtained and cloned into eukaryotic expression vector pCMV-GFP-C to construct the recombinant eukaryotic expression vector pCMV-GFP-C/hSlit2,and the recombinant plasmid was transfected into 293 cells by calcium phosphate coprecipitation. The expression of hSlit2 protein was analyzed by Western blot. Results:The eukaryotic expression recombinant plasmid pCMV-GFP-C/hSlit2 was successfully constructed and confirmed by restriction enzyme digestion,and the expression of hSlit2 cDNA was detected by Western blot. Conclusion:The recombinant eukaryotic expression vector is correctly constructed,and the study offers an experimental base for the study of the fuction of hSlit2 protein and its proteolytic fragments in central nervous system.

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蒋羽清,刘锦波,赵俊丽,张伟锋,夏海滨.人Slit2全长cDNA的真核表达及鉴定[J].南京医科大学学报(自然科学版),2010,(2):183-187

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