Gene transfer into subcultured endometrial cells using lipofection.
Lipofection using the Lipofectin reagent was optimized to transiently transfect subcultured guinea pig endometrial stromal cells with a beta-galactosidase gene driven by a simian virus 40 promoter. Efficient transfection was obtained in the following conditions: a value of six for the ratio of lipofectin to DNA, a low cellular density (10(5) cells per 35-mm well) at the time of subculture (48 h before lipofection) and a lipofection duration of 12 hours. Lipofection was compared to calcium phosphate precipitation previously optimized in the same culture model. At a low cellular density, the lipofection method was found to be more efficient than the calcium phosphate precipitation. This result gives a great relevance to lipofection since the cultured cells available in an experiment are often limited. Then, using cells at low density and a plasmid containing the chloramphenicol acetyltransferase (cat) gene linked to an estrogen response element, it was shown that the lipofection procedure is a suitable tool for the evaluation of gene regulation by estrogen.
Lascombe
I
I
Medical School Hospital Jean Minjoz, Besançon, France.
Mougin
P
P
Vuillermoz
C
C
Adessi
G L
GL
Jouvenot
M
M
D000818:Animals; D002130:Calcium Phosphates; D002478:Cells, Cultured; D004717:Endometrium / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004958:Estradiol / Q000494:pharmacology; D005069:Evaluation Studies as Topic; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D006168:Guinea Pigs; D008081:Liposomes; D010714:Phosphatidylethanolamines; D011960:Receptors, Estrogen / Q000235:genetics; D014162:Transfection / Q000379:methods*; D001616:beta-Galactosidase / Q000235:genetics
D002130:Calcium Phosphates; D008081:Liposomes; D010714:Phosphatidylethanolamines; D011960:Receptors, Estrogen; C485828:alpha-tricalcium phosphate; C485830:tetracalcium phosphate; D004958:Estradiol; C485838:calcium phosphate, monobasic, anhydrous; C020888:1,2-dielaidoylphosphatidylethanolamine; C020243:calcium phosphate; D001616:beta-Galactosidase; C485829:calcium phosphate, dibasic, anhydrous