Fig. 4.

Flow cytometric analysis and estimation of S-phase duration in Arabidopsis and rice suspension cell cultures. Four (Arabidopsis) or three (rice) days after transfer, cells in logarithmic growth phase were pulse-labeled with 10 μM (Arabidopsis) or 25 μM (rice) EdU for 30min and then chased with 100 μM thymidine for select hourly intervals. Cells were pelleted, fixed, washed, and frozen. Nuclei were prepared and analyzed by flow cytometry as described in Fig. 1. (A–D) Bivariate flow cytograms and corresponding histograms of relative DNA content of S-phase nuclei isolated from cell suspension cultures of (A, B) Arabidopsis and (C, D) rice. Data represent single biological replicates. (E, F) RM plots for (E) Arabidopsis and (F) rice cell cultures. For each species, RM values from multiple biological replicates were calculated and plotted as a function of time. Segmented regression analysis was applied to the data of combined biological replicates as described in Fig. 2. The R ² value of the segmented fit, slope of the first component line, and lower and upper average S-phase duration estimates are as follows: (E) Arabidopsis, R ²=0.99, y=0.2326x+0.5654 and 1.5h/1.9h; and (F) rice, R ²=0.95, y=0.0926x+0.5676 and 2.6h/4.7h.