Use of green fluorescent protein variants to monitor gene transfer and expression in mammalian cells

Two mutants of the green fluorescent protein (GFP), RSGFP4 and GFPS65T, have been recently created which differ from the wildtype GFP of A. victoria in their excitation maxima. Here we show that human fibroblasts transfected with either of the two mutant GFP genes emit a green fluorescence that is 1...

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Bibliographic Details
Published in:Nature biotechnology 1996-05, Vol.14 (5), p.606-609
Main Authors: Cheng, Linzhao, Fu, Joan, Tsukamoto, Ann, Hawley, Robert G
Format: Article
Language:English
Subjects:
Aequorea victoria
Agriculture
Animals
Bioinformatics
Biomedical and Life Sciences
Biomedical Engineering/Biotechnology
Biomedicine
Biotechnology
Cell Line
Fluorescence
Gene Expression
Gene Transfer Techniques
Genetic Variation
Genetic Vectors
Green Fluorescent Proteins
Humans
Life Sciences
Luminescent Proteins - genetics
Mice
Mutation
research-article
Retroviridae - genetics
Transduction, Genetic
Transfection
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Summary:Two mutants of the green fluorescent protein (GFP), RSGFP4 and GFPS65T, have been recently created which differ from the wildtype GFP of A. victoria in their excitation maxima. Here we show that human fibroblasts transfected with either of the two mutant GFP genes emit a green fluorescence that is 18-fold brighter than the cells transfected with the wildtype GFP gene. Retroviral vectors expressing the improved GFP gene were also constructed to determine their suitability for stable gene transduction into mammalian cells. The inclusion of the RSGFP4 gene in a retroviral vector did not reduce the viral titer and resulted in a fluorescent signal in viable transduced cells detectable by both fluorescence microscopy and fluorescence-activated cell sorter (FACS) analysis. Therefore, the improved mutant GFP provides a vital marker for monitoring gene transfer and expression in mammalian cells.
ISSN:1087-0156
1546-1696
DOI:10.1038/nbt0596-606