Green fluorescent protein marks skeletal muscle in murine cell lines and zebrafish.
The green fluorescent protein (GFP) acts as a vital dye upon the absorption of blue light. When the gfp gene is expressed in bacteria, flies or nematodes, green fluorescence can be directly observed in the living organism. We inserted the cDNA encoding this 238-amino-acid (aa) jellyfish protein into an expression vector containing the rat myosin light-chain enhancer (MLC-GFP) to evaluate its ability to serve as a muscle-specific marker. Transiently, as well as stably, transfected C2C12 cell lines produced high levels of GFP distributed homogeneously throughout the cytoplasm and was not toxic through several cell passages. Expression of MLC-GFP was strictly muscle-specific, since Cos 7 fibroblasts transfected with MLC-GFP did not fluoresce. When GFP and beta Gal markers were compared, the GFP signal was visible in the cytoplasm of the living cell, whereas visualization of beta Gal required fixation and resulted in deformation of the cells. When the MLC-GFP construct was injected into zebrafish embryos, muscle-specific gfp expression was apparent within 24 h of development. gfp expression was never observed in non-muscle tissues using the MLC-GFP construct. Transgenic fish continued to express high levels of gfp in skeletal muscle at 1.5 months, demonstrating that GFP is an effective marker of muscle cells in vivo.
Moss
J B
JB
Cardiovascular Research Center, Massachusetts General Hospital East, Charlestown 02129, USA. moss@helix.mgh.harvard.edu
Price
A L
AL
Raz
E
E
Driever
W
W
Rosenthal
N
N
D000818:Animals; D030801:Animals, Genetically Modified; D002460:Cell Line; D002461:Cell Line, Transformed; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics*; D015336:Molecular Probe Techniques; D018482:Muscle, Skeletal / Q000378:metabolism*; D018994:Myosin Light Chains / Q000032:analysis / Q000235:genetics; D051381:Rats; D037401:Scyphozoa; D015027:Zebrafish
D008164:Luminescent Proteins; D018994:Myosin Light Chains; D049452:Green Fluorescent Proteins