FLOW CYTOMETRIC AND MICROSCOPIC ANALYSIS OF GFP-TAGGED PSEUDOMONAS-FLUORESCENS BACTERIA

The gene encoding the green fluorescent protein (GFP) from the jellyfi sh Aequorea victoria was assembled into an expression cassette for bac teria by fusing it to the T7 gene10 ribosome binding site and the stro ng, constitutive promoter PpsbA from Amaranthus hybridus. By using Tn5 -based transposon delivery systems, Pseudomanas fluorescens bacteria w ere chromosomally tagged with gfp. We demonstrate that expression of a single copy gfp gene is sufficient to permit the visualization of bac teria by epifluorescence and laser confocal microscopy and detection b y flow cytometry. The green fluorescent phenotype was detectable in al l growth phases even under nutrient-limited conditions.