title: Transfection of cells with DNA encoding a visible fluorescent protein-tagged lipid-binding domain.
abstract: Visualization of a certain lipid species can be achieved by producing a fusion protein between a lipid-binding domain and a visible fluorescent protein (VFP). After a DNA construct for a VFP-tagged lipid-binding domain has been prepared, the desired cell line is transfected with the DNA and visualized using fluorescence microscopy, as described here. The DNA encoding a VFP-tagged lipid-binding domain is isolated from Escherichia coli, and the cells to be transfected are grown on glass-bottom dishes or on coverslips. We routinely perform transfection with commercially available reagents, although, depending on the cell line, the calcium phosphate precipitation method may provide an economic alternative.
authors.forename: Carsten
authors.initials: C
authors.lastname: Schultz
authors.forename: Anne B
authors.initials: AB
authors.lastname: Neef
authors.forename: Theodorus W
authors.initials: TW
authors.lastname: Gadella
authors.forename: Joachim
authors.initials: J
authors.lastname: Goedhart
mesh_terms: D002460:Cell Line; D004247:DNA / Q000378:metabolism*; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D014162:Transfection / Q000379:methods*
chemical_list: D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D004247:DNA