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Published on: June 19, 2013
[Construction of a recombinant lentiviral expression vector carrying carcinoembryonic antigen gene and its expression
Hong Zheng1, Wen-ling Zhang, Guo-qiang Zhao
1Department of Pathologic Physiology, School of Basic Medical Science, Zhengzhou University, Zhengzhou, China. zhhong@henu.edu.cn
Objective:
To construct a lentiviral expression vector of human carcinoembryonic antigen (CEA), and identify its expression in dendritic cells (DCs).
Methods:
Human CEA-encoding sequence was amplified, purified, ligated with lentiviral vector plasmid pLentiGFP and verified by sequencing. The verified recombinant vector plasmid (pLentiGFP-CEA), the packaging plasmid p 8.2 and pVSV-G were transfected into 293T cells by Lipofectamine(TM) 2000 reagent. The supernatant of the cultured 293T cells was collected to infect the DCs. The expression of CEA in the transfected DCs was assayed by RT-PCR and Western blotting.
Results:
CEA lentiviral vector was highly expressed in the transfected DCs as observed using fluorescence microscope 48 h after the the transfection. The human CEA gene was successfully amplified by RT-PCR with a length of about 2100 bp. Western blotting also showed CEA expression in the transfected DCs.
Conclusion:
The human CEA lentiviral expression vector has been successfully constructed and the functional CEA protein can be expression in the transfected DCs. This facilitates further studies of the function of CEA at the molecular level.
