Influential factors on the efficiency in transfecting human keratinocytes with plasmid-liposome complexes / 基础医学与临床
Basic & Clinical Medicine
;
(12): 180-184, 2001.
Article
in Chinese
| WPRIM
| ID: wpr-411146
ABSTRACT
To investigate the optimum transfection condition in transfecting human keratinocytes with plasmid-liposome complexes in vitro,the cultured human keratinocytes at 60% ~ 100% confluences were exposed to the eukaryotic expression plasmid,pCMV*SPORT-β-gal,coated with LipofectAMINE in different DNA/liposome mixing concentration ratios.After cultured for another 48 hours following the ends of 6 ~ 24 hours exposures to the DNA-liposome complexes,the transfected human keratinocytes were visualized by β-galactosidase staining.Then,the transfection efficiency was determined by calculating the rate of β-galactosidase staining positive cells.β-galactosidase expression was showed clearly in human keratinocytes transfected with the DNA-liposome complexes.The highest efficiency was achieved with cultured cells at 80% and 90% confluences,demonstrating by the transfection rates of (31.35±1.35)% and (32.32±2.47)% respectively.Meanwhile,the essential transfection conditions for these efficiencies were in coating pCMV*SPORT-β-gal DNA of 1.5μg/100μL with LipofectAMINE of 12.5μL/100μL,and exposing the cells to the DNA-liposome complexes for 8 hours.These results indicate that LipofectAMINE could effectively transfer eukaryotic expression plasmid into human keratinocytes in vitro,for which the optimization of transfection conditions involve in cells growth state,DNA/liposome mixing concentration ratio,and exposure time of cells to DNA-liposome complexes.
Full text:
Available
Index:
WPRIM (Western Pacific)
Language:
Chinese
Journal:
Basic & Clinical Medicine
Year:
2001
Type:
Article
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