Subpopulation tracking by volume histograms.

<p>Continuous volume measurements of an arrest in <i>cdc28-as1</i> yeast over a 3-hour timecourse show cells arrested at various points in the cell cycle. (<b>A</b>) Histogram features, such as bounds, on single-mode volume distributions separately analyze large and small cells. (<b>B</b>) At room temperature (∼22°C), cell volume steadily increases with a single population mode, and (<b>C</b>) the arrested cells' volume growth rates increase with cell size. Error bars on <i>cdc28-as1</i> data indicate the 95% confidence interval for the linear regression. The growth rate average (n = 6) and measured standard deviation (error bar) for formaldehyde-fixed cells with a 60 min window are shown for comparison to measurement error. (<b>D</b>) Subpopulations emerge at the optimal temperature for yeast (∼30°C) and a later timepoint (∼120 min), and the mode of each subpopulation is identified and tracked. These subpopulations likely represent slow-growing (SG), G1-arrested (G1), and metaphase-arrested (M) cells. (<b>E</b>) Histogram features are determined for each 150 s file, similar to those in <b>B</b>. The color of the solid circles indicates the fraction of cells in each subpopulation. Growth rates calculated with data in <b>B</b> and <b>E</b> are by linear regression on a 60 or 180 min window, as indicated. (<b>F</b>) Growth rates are separately calculated for a one hour window before and after the emergence of subpopulations. Fixed cell data is identical to that in <b>C</b>. As observed for room temperature measurements, growth rates are related to cell size and likely a result of the multi-phase arrest.