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MOLECULAR CLONING, EXPRESSION AND SUBCELLULAR LOCALIZATION OF HUMAN OCT-4 PROTEIN

MOLECULAR CLONING, EXPRESSION AND SUBCELLULAR LOCALIZATION OF HUMAN OCT-4 PROTEIN
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摘要 Objective To clone, express, purify human Oct-4 and detect its subcellular localization. Methods Human Oct-4 cDNA was amplified by RT-PCR strategy. Oct-4 protein was induced and expressed in BL21(DE3) strain. Furthermore, the protein was purified with Ni-NTA resin. Subsequently, site-directed mutagen-esis of Oct-4 (aa 236 - 240) was introduced. Finally, the subcellular localization of wide type Oct-4 and mutant Oct-4 was examined by immunofluorescent cytochemistry staining and confocal laser scanning microscope analysis. Results The full length cDNA of human Oct-4 was 1083 bp. Human Oct-4 encoded a 55 kd protein by prokaryotic vector in E coli. Compared with pure nuclear localization of wide type Oct-4, mutant Oct-4 was mostly enriched in the cytoplasm. Conclusion The cloning, expression and investigation of subcellular localization of human Oct-4 are basis of studying its biological function. Objective To clone, express, purify human Oct-4 and detect its subcellular localization. Methods Human Oct-4 cDNA was amplified by RT-PCR strategy. Oct-4 protein was induced and expressed in BL21 （DE3） strain. Furthermore, the protein was purified with Ni-NTA resin. Subsequently, site-directed mutagenesis of Oct-4 （aa 236 -240 ） was introduced. Finally, the subcellular localization of wide type Oct-4 and mutant Oct-4 was examined by immunofluorescent cytochemistry staining and confocal laser scanning microscope analysis. Results The full length cDNA of human Oct-4 was 1083bp. Human Oct-4 encoded a 55 kd protein by prokaryotic vector in E coli. Compared with pure nuclear localization of wide type Oct-4, mutant Oct-4 was mostly enriched in the cytoplasm. Conclusion The cloning, expression and investigation of subcellular localization of human Oct-4 are basis of studying its biological function.
作者 李香群 金颖
机构地区 Department of Developmental Biology School of Basic Medicine
出处 《Journal of Shanghai Second Medical University(Foreign Language Edition)》 2006年第1期15-19,25,共6页 上海第二医科大学学报（英文版）
关键词 Oct-4 pluripotency nuclear localization signal 分子克隆 基因表达 亚细胞 OCT-4蛋白
分类号 Q78 [生物学—分子生物学]
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参考文献10

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二级参考文献53

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共引文献37

Journal of Shanghai Second Medical University(Foreign Language Edition)
2006年 第1期

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