Abstract: Objective: To construct the eukaryotic expression vectorsof human Aurora A(AURKA), and to detect the expression and purification of AURKA in Pichia pastoris X33 and MCF-7 cells. Methods: The AURKA was amplified by polymerase chain reaction (PCR). The genes were digested with XhoⅠ and XbaⅠ, and respectively ligated into pPICZαA and pcDNA3.1(+) to construct the recombinant plasmids pPICZαA- AURKA and pcDNA3.1(+)-AURKA which were transfected into Pichia pastoris X33 and MCF-7 cells by Gene Pulser Xcell^TM Electroporation System. The expressions of AURKA protein were detected by SDS-PAGE and Western blotting methods after screening in Pichia pastoris X33 and MCF-7 cells, respectively. The purification of proteins was determined using Ni-NTA column. The activity of AURKA was analyzed by autoradiography. The proliferation of MCF-7 cells after transfected with AURKA was detected by cell counting method. Results: The 1 212 bp genes were amplified from the pPICZαA-AURKA and pcDNA3.1(+)-AURKA by PCR, which indicated that two eukaryotic expression vectors were successfully constructed. The relative molecular mass of recombinant AURKA protein was approximately 50 000. The recombinant AURKA phosphorylated the histone H3 by autoradiography, which manifested that it had activity. Compared with the cells transfected with empty plasmid and WT MCF-7 cells, the number of alive MCF-7 cells was significantly increased after transfected with AURKA for 24 and 48 h. Conclusion: AURKA is successfully expressed and purified in Pichia pastoris X33 and MCF-7 cells. The over-expression of AURKA can promote the proliferation of MCF-7 cells

Key words: human Aurora A, MCF-7 cells, cell proliferation, Pichia pastoris X33