Table 1.
Bacterial strains and plasmids.
Fig 1.
Fluorescence spectra of GFP, E2-Orange and mCherry.
Shown are normalized excitation and emission (A) and synchronous (B) fluorescence spectra of E. coli cultures expressing GFP, E2-Orange and mCherry.
Table 2.
Properties of fluorescent proteins in vitro and in living E. coli cells.
Fig 2.
Synchronous spectra acquisition and CP decomposition of fluorescence from mixtures of four E. coli TOP10 strains expressing different FPs constitutively.
(A) Typical SFS dataset of FP labeled strains mixtures (excerpt of 11 spectra out of 35). (B-D) Outcomes of CP decomposition. The fluorescence profiles estimated from CP analysis of mixtures of FP labeled strains (solid lines) are compared to those obtained from single labeled strains benchmarks (dashed lines). (B) Fluorescence profile as a function of labeled strain concentration (dilution level). (C) Synchronous fluorescent spectra. (D) Fluorescence profile as a function of strains mixing ratio. The mixture pattern for this experiment is presented in S1 Fig. YFP: TurboYFP; E2O: E2-Orange; DsRed: DsRed-express2.
Fig 3.
Spectral decomposition of fluorescence from mixtures of two P. aeruginosa PAO1 iron bioreporter strains harboring pvdA-dsred-express2 and bfrB-e2-orange fusions.
Shown are (A) the spectra of the four fluorescent sources identified from CP analysis and their profile as a function of (B) iron concentration and (C) the ratio of bioreporter strains in the mixture. The mixture pattern for this experiment is presented in S2 Fig. (D) Synchronous spectra of purified pyoverdine (solid line) and cultures of PAO1 wild type (WT, dashed line) and PAO1 ∆pvdA (large dashed lines) cells grown in low-iron DCAA medium. The later spectrum was normalized with respect to that obtained for PAO1 wild type. (E) Synchronous spectra of WT PAO1 culture supernatant as a function of pH. The DCAA growth medium supernatant was diluted tenfold in different buffers with pH adjusted to 5.2 and 6.2 (40 mM MES), 7.4 (40 mM MOPS) and 8, 9 and 10 (40 mM Tris-HCl).
Fig 4.
Spectral decomposition of fluorescence from mixtures of three P. aeruginosa PAO1 iron bioreporter strains harboring pvdS-gfp, pvdA-dsred-express2 and bfrB-e2-orange fusions.
(A) Raw synchronous spectra of strain mixtures incubated with 0.3, 1 and 3 μM FeCl3 showing the relative intensity of fluorescent signals. (B) Spectra of the four fluorescent sources identified from CP analysis performed independently on 400–470 nm and 470–600 nm wavelength ranges. (C) Profile of fluorescence sources as a function of iron concentration. (D) Profile of fluorescence sources as a function of the ratio of bfrB-e2-orange reporter strain in the mixture. The mixture pattern for this experiment is presented in S3 Fig. PVD: pyoverdine; af-PVD and bf-PVD: “acid” and “basic” forms of pyoverdine.