Green fluorescent protein and its derivatives as versatile markers for gene expression in living Drosophila melanogaster, plant and mammalian cells.
We have investigated the utility of the green fluorescent protein (GFP) as a marker for gene expression in living adult Drosophila melanogaster (Dm) and cultured plant and mammalian cells. Using Dm, we generated transgenic flies bearing a glass-responsive gfp fusion gene to test the utility of GFP as a spatial reporter. In the adult living fly, GFP is clearly visible in the ocelli and the eye. We have optimized the use of filters for distinguishing the GFP signal from abundant autofluorescence in living Dm. In addition, we have used GFP to identify photoreceptor cells in pupal eye cultures that have been fixed and stained according to standard histological procedures. GFP was also detected in individual living plant cells following transient transfection of soybean suspension cultures, demonstrating that GFP is an effective transformation marker in plant cells. Similarly, transient transfection of mammalian cells with a modified form of GFP, S65T, allowed detection of single living cells expressing the reporter. This modified form of GFP gave a robust signal that was resistant to photobleaching. We then used a CellScan system exhaustive photon reassignment (EPR) deconvolution algorithm to generate high-resolution three-dimensional images of GFP fluorescence in the living cell.
Plautz
J D
JD
NSF Center for Biological Timing, University of Virginia, Charlottesville 22903, USA.
Day
R N
RN
Dailey
G M
GM
Welsh
S B
SB
Hall
J C
JC
Halpain
S
S
Kay
S A
SA
D000818:Animals; D030801:Animals, Genetically Modified; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000235:genetics*; D015870:Gene Expression; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D037401:Scyphozoa; D013025:Glycine max / Q000166:cytology; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D008164:Luminescent Proteins; C087289:gl protein, Drosophila; D049452:Green Fluorescent Proteins