[Construction of recombinant plasmid pEGFP-N1/MnSOD and its express in vitro].
OBJECTIVE
To construct eukaryotic expression plasmid of rat Mn-superoxide dismutase (MnSOD) gene pEGFP-N1/MnSOD and express it in primary culture system of marginal cells (MC) of the rats.


METHODS
The aimed segments were obtained from rat myocardial tissue and were inserted into a eukaryotic expression plasmid pEGFP-N1/MnSOD. The recombinant expression plasmid pEGFP-N1/MnSOD was transfected into MC by lipofectamine 2000-mediated gene transfer method, which were observed through co-Focus Fluorescence microscopy. The percentage of transfection was determined by fluorescence activated call sorting (FACS) analysis.


RESULTS
Rat MnSOD cDNA eukaryotic expression plasmid pEGFP-N1/MnSOD have been successfully constructed. Green fluorescent protein(GFP) and MnSOD protein could be contacted in the transfected MC 48 hours after transfection. The percentage of transfection was 23.47%.


CONCLUSIONS
Rat MnSOD cDNA eukaryotic expression plasmid pEGFP-N1/MnSOD have been successfully constructed and expressed successfully in MC. The research paved the way for antioxidant gene therapy of inner ear.
Yang
Yang
Y
Department of Otolaryngology, the Union Hospital, Tongjing Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
Kong
Weijia
W
D000818:Animals; D015870:Gene Expression; D005822:Genetic Vectors; D010957:Plasmids; D051381:Rats; D017207:Rats, Sprague-Dawley; D013316:Stria Vascularis / Q000166:cytology*; D013482:Superoxide Dismutase / Q000235:genetics*; D014162:Transfection
D013482:Superoxide Dismutase