Construction of eukaryotic expression vector pcDNA3-MAGE-1 and its stable expression in mouse Hepa1-6 cells

Objective To construct the eukaryotic expression vector and express in mouse liver cancer cell Hepa1-6,and observe reproductive activity and tumorigenesis ability of transfected cells.Methods The MAGE-1 total length sequence was amplified from SMMC-7721 by PCR and linked to pcDNA3 to construct pcDNA3-MAGE-1.The expression vector was transfected into Hepa1-6 cells by lipofectamine,and then the positive clones were screened by G418.The expressions of mRNA and protein in positive clones were detected by RT-PCR and Western blotting.Cell growth and proliferation in Hepa1-6 cells and Hepa1-6-MAGE-1 cells were detected by MTT,then they were inoculated in the right back subcutaneous of mice.Results pcDNA3-MAGE-1 was constructed and transfected into Hepa1-6 cells,the expressions of MAGE-1 mRNA and protein were detected in Hepa1-6-MAGE-1 cells by RT-PCR and Western blotting,there was no statistical difference in cell proliferation between two groups.The mice were all infected with tumor in two groups,and there was no difference in tumor size.Conclusion The eukaryotic expression vector pcDNA3-MAGE-1 is successfully constructed,and the Hepa1-6 cell line which can stably express human MAGE-1 gene is established with good proliferation and tumorigenesis ability.