Construction of different mutants of HA-tagged human RAGE gene and their eukaryotic expression.
- Author:
Wei-wei CHENG
1
;
Yu-sheng LI
;
Xiao-wei GONG
;
Lin-lin ZHAO
;
Ji-gang WANG
;
Peng DENG
;
Yong JIANG
Author Information
- Publication Type:Journal Article
- MeSH: Cell Line; Cloning, Molecular; Eukaryotic Cells; metabolism; Genetic Vectors; genetics; Humans; Mutagenesis, Site-Directed; Mutation; Receptor for Advanced Glycation End Products; Receptors, Immunologic; biosynthesis; genetics
- From: Journal of Southern Medical University 2008;28(10):1779-1781
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct eukaryotic expression vectors for HA-tagged receptor for advanced glycation end products (RAGE) mutants.
METHODSSite-directed mutagenesis was applied to wild-type RAGE gene cloned in the pcDNA3 vector with HA tag to obtain the mutants pcDNA3-HA-RAGE(S391A), pcDNA3-HA-RAGE(S399A), pcDNA3-HA-RAGE(S400A), and pcDNA3-HA-RAGE(T401A). After identification by sequencing, the mutants were transfected into HEK293 cells, and the expression of these mutants were detected by Western blotting using anti-HA antibody.
RESULTSThe HA-tagged RAGE mutants constructed were verified successfully by sequencing, and highly expressed in HEK293 cells.
CONCLUSIONThe success in constructing HA-tagged RAGE mutants, which are highly expressed in eukaryotic cells, may facilitate the functional study of RAGE in cell signal transduction.
