Construction of pEGFP-N1-Fcy::Fur recombinant plasmid and observation of its expression in ovarian cancer cells
- VernacularTitle:pEGFP-N1-Fcy::Fur重组质粒的构建及其在卵巢癌细胞中的表达
- Author:
Xia WU
- Publication Type:Journal Article
- Keywords:
Suicide gene;
Fcy:Fur;
Green fluorescent protein;
Gene transfection;
Eukaryotic expression plasmid
- From:
Journal of Chongqing Medical University
2007;0(11):-
- CountryChina
- Language:Chinese
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Abstract:
Objective:To construct a EGFP(Enhanced green fluorescent protein)-labled euk-aryotic expression plasmid of Fcy::Fur suicide gene and to detect its expression in SKOV3 cell line. Methods:With the technology of gene re-arrangement,Fcy::Fur gene in pORF-Fcy:Fur plasmid was subcloned into pEGFP-N1 vector,with its correctness evaluated by the means of r-estriction enzyme analysis and sequencing.It was transfected into SKOV3 cells with lipofectin,the transient expression of GFP was observed under flu- orescence microscope after 24 hours and detected by Western blot. Results:Correct construction of pEGFP-N1-Fcy::Fur was identi- fied by methods of restriction enzyme analysis and nucleotide sequence determination.A total of 60% transfe-cted cells emitted out green fluorescence under fluorescent microscope after 24 h after transfecti-.on. Fcy::Fur gene expressed by the transfected cells were testified by Western blot. Conclusion:The recombinant eukaryotic expression vectors have been constructed successfully and effec- tive-ly expressed in ovarian cancer cells,which may provide an experimental basis for gene therapy of ovarian cancer.
