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Cold Spring Harbor Protocols, 7(2010), p. pdb.prot5457-pdb.prot5457

DOI: 10.1101/pdb.prot5457

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Transfection of Cells with DNA Encoding a Visible Fluorescent Protein-Tagged Lipid-Binding Domain

Journal article published in 2010 by Carsten Schultz, Anne B. Neef, Theodorus W. Gadella ORCID, Joachim Goedhart ORCID
This paper is made freely available by the publisher.
This paper is made freely available by the publisher.

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Abstract

INTRODUCTIONVisualization 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.