Gene transfer into established and primary fibroblast cell lines: comparison of transfection methods and promoters.
The stable transfection of immortalized Rat-1 and rat skin primary fibroblast cell lines by calcium phosphate precipitation, lipofection and electroporation methods have been examined. The lipofection method was found to be better than the other methods in terms of higher transfection efficiency and convenient use. Expression of beta-galactosidase from two different viral promoters showed that the level of transgene expression depends on the promoter strength in a particular cell type. The results presented here show that the transgene expression is extremely variable among different colonies generated from individually transfected cells. Therefore, it is necessary to examine individual colonies of cells for the production of reporter gene to obtain cell lines expressing high amounts of gene products.
Ray
J
J
Department of Neurosciences, University of California, San Diego La Jolla 92093-0627.
Gage
F H
FH
D000818:Animals; D001358:Avian Sarcoma Viruses; D002130:Calcium Phosphates; D002999:Clone Cells; D004351:Drug Resistance / Q000235:genetics; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D008979:Moloney murine leukemia virus; D009355:Neomycin; D011401:Promoter Regions, Genetic / Q000502:physiology; D051381:Rats; D011916:Rats, Inbred F344; D013539:Simian virus 40; D012867:Skin / Q000166:cytology; D014162:Transfection / Q000379:methods*; D001616:beta-Galactosidase / Q000096:biosynthesis
D002130:Calcium Phosphates; C485828:alpha-tricalcium phosphate; C485830:tetracalcium phosphate; C485838:calcium phosphate, monobasic, anhydrous; C020243:calcium phosphate; D001616:beta-Galactosidase; D009355:Neomycin; C485829:calcium phosphate, dibasic, anhydrous