Construction of a recombinant adenovirus expression vector for human renal tumor- associated antigen G250 gene with AdMax system.
- Author:
Huan QI
1
Author Information
- Publication Type:Journal Article
- MeSH: Adenoviridae; genetics; Antigens, Neoplasm; biosynthesis; genetics; Carcinoma, Renal Cell; genetics; immunology; pathology; Dendritic Cells; metabolism; Genetic Vectors; genetics; Humans; Kidney Neoplasms; genetics; immunology; pathology; RNA, Messenger; biosynthesis; genetics; Recombinant Proteins; biosynthesis; genetics; Transfection; Tumor Cells, Cultured
- From: Journal of Southern Medical University 2008;28(9):1617-1625
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct a adevoviral vector harboring human renal tumor-associated antigen G250 gene for transfecting the dendritic cells (DCs) and treating renal tumors.
METHODSThe G250 genes were cloned into the shuttle plasmid pDC316 to construct pDC316-G250, which was cotransfected with the rescue plasmid pBHGlox(delta)E1,3Cre into 293 cells to obtain the recombinant adenovirus Ad/G250. The inserted gene and its expression were identified by RT-PCR and fluorescence-activated cell sorting (FACS) after recombinant adenovirus transfection of the DCs. The recombinant adenoviral vector was purified by CsCl banding and titrated by TCID50.
RESULTS AND CONCLUSIONThe recombinant adenoviral vector of G250 gene was successfully constructed and high titer of the recombinant adenoviruse was obtained. G250 mRNA and protein expressions were identified in Ad/G250-transfected DCs. The titer of the virus stocks reached 5.6x10(9) IU/ml.
