Grb2在MCF-7 乳腺癌细胞中的核定位研究

目的　构建Grb2的红色荧光蛋白融合表达质粒pDsRed1 C/Grb2 ,并观察Grb2在MCF 7乳腺癌细胞的核定位情况。方法　用PCR方法扩增Grb2cDNA ,与红色荧光蛋白表达质粒pDsRed1 C重组构建融合蛋白表达质粒pDsRed1 C/Grb2 ,酶切、测序鉴定后 ,瞬时转染MCF 7乳腺癌细胞株 ,用荧光显微镜观察Grb2的核定位情况。结果　pDsRed1 C/Grb2质粒有正确的阅读框 ,融合表达蛋白DsRed1 C/Grb2可定位于MCF 7乳腺癌细胞核内。结论　成功构建了Grb2的红色荧光蛋白融合表达质粒pDsRed1 C/Grb2 ,在MCF 7乳腺癌细胞株中 ,Grb2除存在于胞浆外 ,也可定位于细胞核内 ,可能与其参与核内基因的转录调控有关。

Nucleic localization of Grb2 in MCF-7 cells

Objective To construct the recombinant plasmid pDsRed1-C/Grb2 expressing a fusion construction of red fluorescent protein pDsRed1-C and Grb2 and observe the nucleic localization of Grb2 in MCF-7 cells. Methods A cDNA fragment encoding Grb2 was amplified by PCR and inserted into MCS of red fluorescent protein pDsRed1-C. After identification of the positive clone by digestion of restriction endonuclease and sequencing, the recombinant construct was transiently transfected into MCF-7 cells and the nucleic localization of the fusion protein DsRed1-C/Grb2 was observed by fluorescence microscopy. Results The fusion protein, expressed by the plasmid with a correct reading frame, could be localized in the nuclei of MCF-7 cells. Conclusion The recombinant plasmid pDsRed1-C/Grb2, which encodes fusion protein of DsRed1-C and Grb2, has been successfully constructed. Grb2 can be localized in the nuclei of MCF-7 cells except as a cytosolic protein.