Citation

Wu X. Journal of Chongqing Medical University 2007; (12).

Copyright

(Copyright © 2007)

DOI

unavailable

PMID

unavailable

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.

Language: zh

Keywords

Suicide gene; Eukaryotic expression plasmid; Fcy:Fur; Gene transfection; Green fluorescent protein