Recombinant plasmid construction and fusion protein expression and localization of human ER αgene

Objective To construct recombinant expression plasmid of human estrogen recptor α(hERα) gene and identify its protein expression and localization. Methods The hERα coding sequence was amplified by polymerase chain reaction(PCR) method and subcloned into pEGFP-C1 vector. After the target region was sequenced,the plasmid was transfected into MCF-7 cell lines. The expression of the recombinant plasmid in breast cancer cells(MCF-7 cells) was proved by Western blot and the localization of pEGFP-hERα C was observed by using laser scanning confocal microscopy. Results HERαwas constructed into expressing vector pEGFP-C1 successfully,the length of the fragment was 1800bp identified by restriction enzymes digestion. The expression of pEGFP-hERα fusion protein was detected by Western blot in MCF-7 cells,with a molecular weight 95KD,and localized in the nucleus of MCF-7 cells. Conclusion The recombinant plasmid of pEGFP-hERαwas constructed successfully and the fusion protein was localized in nucleus of MCF-7 cells.