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EGFP and DsRed expressing cultures of Escherichia coli imaged by confocal, two‐photon and fluorescence lifetime microscopy
Author(s) -
Jakobs Stefan,
Subramaniam Vinod,
Schönle Andreas,
Jovin Thomas M.,
Hell Stefan W.
Publication year - 2000
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(00)01896-2
Subject(s) - green fluorescent protein , fluorescence , escherichia coli , confocal microscopy , förster resonance energy transfer , fluorescence microscope , microscopy , two photon excitation microscopy , fluorescence lifetime imaging microscopy , fluorescent protein , biophysics , confocal , protein tag , chemistry , microbiology and biotechnology , biology , recombinant dna , gene , biochemistry , fusion protein , optics , physics
The green fluorescent protein (GFP) has become an invaluable marker for monitoring protein localization and gene expression in vivo. Recently a new red fluorescent protein (drFP583 or DsRed), isolated from tropical corals, has been described [Matz, M.V. et al. (1999) Nature Biotech. 17, 969–973]. With emission maxima at 509 and 583 nm respectively, EGFP and DsRed are suited for almost crossover free dual color labeling upon simultaneous excitation. We imaged mixed populations of Escherichia coli expressing either EGFP or DsRed by one‐photon confocal and by two‐photon microscopy. Both excitation modes proved to be suitable for imaging cells expressing either of the fluorescent proteins. DsRed had an extended maturation time and E. coli expressing this fluorescent protein were significantly smaller than those expressing EGFP. In aging bacterial cultures DsRed appeared to aggregate within the cells, accompanied by a strong reduction in its fluorescence lifetime as determined by fluorescence lifetime imaging microscopy.