Figure S2.
Lower separase activity reduces centromere separation synchrony and speed. (A and B) Cyan: Frequency distributions and Gaussian fit (continuous lines) of the time difference between the separation of centromeres 1 and 2 or centromeres 2 and 3 for cells carrying the temperature-sensitive separase allele cut1-206, grown in rich medium and then imaged in minimal medium (A) or rich medium (B). The fitted Gaussian distributions of WT cells (black) are shown for comparison. Mean ± SD of the fit; n = number of cells; P values from a two-sample Kolmogorov–Smirnov test. Combined separase mutant datasets from A and B are shown in Fig. 2 A, as distributions were not significantly different (Table S1). (C and D) Sister centromere distance and corresponding kymographs for cut1-206 cells with cen1-tdTomato and cen2-GFP markers. Examples for asynchronous (C) and more synchronous separation (D). (E) Distance between each sister centromere pair in WT and separase-mutant cells (same cells as shown in Fig. 2 B). Distances are aligned to sister chromatid separation (SCS) of the respective chromosome at t = 0. Mean (line) ± SD (shaded area) of the cell population; n = number of cells. Refer to the image caption for details. Panel A shows histograms and Gaussian fits of time differences between centromere separations in separase mutant cells, compared with wild-type distributions, after a shift from rich to minimal medium for imaging. Panel B shows histograms and Gaussian fits of time differences between centromere separations in separase mutant cells, compared with wild-type distributions, when cells were grown and imaged in rich medium. Panel C shows a line graph and kymograph illustrating asynchronous sister centromere separation dynamics in separase mutant cells labeled with cen1-tdTomato and cen2-GFP markers. Panel D shows a line graph and kymograph illustrating more synchronous sister centromere separation dynamics in separase mutant cells labeled with cen1-tdTomato and cen2-GFP markers. Panel E shows line graphs comparing sister centromere distances over time in wild-type and separase mutant cells, aligned to chromosome-specific sister chromatid separation.

Lower separase activity reduces centromere separation synchrony and speed. (A and B) Cyan: Frequency distributions and Gaussian fit (continuous lines) of the time difference between the separation of centromeres 1 and 2 or centromeres 2 and 3 for cells carrying the temperature-sensitive separase allele cut1-206, grown in rich medium and then imaged in minimal medium (A) or rich medium (B). The fitted Gaussian distributions of WT cells (black) are shown for comparison. Mean ± SD of the fit; n = number of cells; P values from a two-sample Kolmogorov–Smirnov test. Combined separase mutant datasets from A and B are shown in Fig. 2 A, as distributions were not significantly different (Table S1). (C and D) Sister centromere distance and corresponding kymographs for cut1-206 cells with cen1-tdTomato and cen2-GFP markers. Examples for asynchronous (C) and more synchronous separation (D). (E) Distance between each sister centromere pair in WT and separase-mutant cells (same cells as shown in Fig. 2 B). Distances are aligned to sister chromatid separation (SCS) of the respective chromosome at t = 0. Mean (line) ± SD (shaded area) of the cell population; n = number of cells.

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