Heterogeneity of cells within exponentially growing populations was addressed in a bacterium, the facultative methylotroph Methylobacterium extorquens AM1. A transcriptional fusion between a well-characterized methanol-inducible promoter (P(mxaF)) and gfp(uv) was used with flow cytometry to analyse the distribution of gene expression in populations grown on either succinate or methanol, correlated with forward scatter as a measure of cell size. These cell populations were found to consist of three major subpopulations defined by cells that were actively growing and dividing, newly divided, and non-dividing. Through the use of flow cytometry, it was demonstrated that a significant percentage of the total population did not respond to carbon shift. In addition, these experiments demonstrated that a small subset of the total population was significantly brighter than the rest of the population and dominated fluorimetry data. These results were corroborated with a continuous flow-through system and laser scanning microscopy, confirming that subpopulations, not discernible in the population average, dominate population response. These results demonstrate that the combination of flow cytometry and microscopic single-cell analysis can be effectively used to determine the dynamics of subpopulations in population response. In addition, they support the concept that physiological diversity in isogenic populations can poise some proportion of the population to respond appropriately to changing conditions.
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