Construction of different mutants of HA-tagged human RAGE gene and their eukaryotic expression / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To construct eukaryotic expression vectors for HA-tagged receptor for advanced glycation end products (RAGE) mutants.</p><p><b>METHODS</b>Site-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.</p><p><b>RESULTS</b>The HA-tagged RAGE mutants constructed were verified successfully by sequencing, and highly expressed in HEK293 cells.</p><p><b>CONCLUSION</b>The 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.</p>