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Published on: February 22, 2015
[Construction of the eukaryotic expression vector pIRES2-EGFP-Axin and its expression in glioma cells]
Li-ying Zhang1, Qing Li, Guang-sheng Chen
1Department of Pathology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China. zly79921@sohu.com
Aim:
To construct the eukaryotic expression vector pIRES2-EGFP-Axin, and to express Axin in C6 glioma cells.
Methods:
The Axin gene was amplified by PCR using pCMV5-HA-Axin as a template, and confirmed by DNA sequencing. The eukaryotic expression vector pIRES2-EGFP-Axin was constructed by introducing Axin DNA fragment into the sites of Nhe I and Sal I of pIRES2-EGFP vector. The plasmid was transfected into the C6 cells using lipofectamine. The expressed EGFP was observed under fluorescent microscope and the Axin protein expression was detected by immunostaining using anti-Axin antibody.
Results:
The eukaryotic expression vector pIRES2-EGFP-Axin was constructed and transfected successfully into C6 glioma cells. The green fluorescence of EGFP was observed in the plasma and nuclei of transfected cells, and Axin protein was only found in the plasma.
Conclusion:
The recombinant expression vector pIRES2-EGFP-Axin was constructed, and the EGFP and Axin gene could be co-expressed in the C6 cells. This study laid a foundation for the further research of the function of Axin in cell differentiation, growth and tumorigenesis.
