title: Transfection and promoter analysis in embryonic stem cells.
abstract: Embryonic stem (ES) cells are useful for the molecular analysis of developmental pathways when combined with efficient genetic manipulation and in vitro differentiation procedures. To facilitate the genetic modification of ES cells, we describe how to analyze the transcriptional activities of different promoter systems. The DNA segments from a number of promoters are subcloned into the multiple cloning sites of the green fluorescent protein (GFP) reporter plasmid. The resulting recombinant plasmids are used to transfect ES cells by lipofection. Using luciferase-normalized GFP assays, we observed differing promoter activities in transient transfections of ES cells. The judicious selection of the optimum promoter system for specific genetic modification of ES cells will ensure the successful generation of transgenic ES cell lines.
authors.affiliation: Molecular Neurobiology Laboratory, McLean Hospital, Harvard Medical School, Belmont, MA, USA.
authors.forename: Sangmi
authors.initials: S
authors.lastname: Chung
authors.forename: Kwang-Soo
authors.initials: KS
authors.lastname: Kim
mesh_terms: D000818:Animals; D004622:Embryo, Mammalian / Q000166:cytology*; D005456:Fluorescent Dyes; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics; D008156:Luciferases; D051379:Mice; D039904:Pluripotent Stem Cells / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics; D014162:Transfection
chemical_list: D005456:Fluorescent Dyes; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D008156:Luciferases