Figure 2
The spindle assembly checkpoint of rad26Δ cells appears functional. A. rad26Δ cells delayed mitosis during TBZ treatment. Cultures of TW1261 (cdc25.22 rad26+) and TW1262 (cdc25.22 rad26Δ) were shifted to 37°C for 4 hours to arrest cells in G2. TBZ (20 μg/ml) was added to respective cultures 30 minutes before shifting to 30°C and releasing into mitosis. Cells were fixed with paraformaldehyde and stained with Calcofluor to visualize septa. B. Chromosome stability was not affected in TBZ-treated rad26Δ cells. The adenine-marked minichromosome of TE787 (rad3Δ), TW1222 (wild type) and TW1224 (rad26Δ) was used to assay chromosome loss following 8 h of TBZ treatment. Bars = Std dev C. Chromosome separation was restrained in rad26Δ cells during TBZ treatment. Cultures of TW1261 and TW1262 were shifted to 37°C for 4 hours and arrested in G2. TBZ was added to respective cultures 30 minutes before shifting to 26°C and releasing into mitosis. Cells were fixed in methanol; chromosome separation was monitored using the Cen1-GFP marker. D. The septation index of asynchronous rad26Δ cultures is elevated. Asynchronous cultures of rad26+ (TE236) and rad26Δ (TE257) cells were split and left untreated or treated with 20 μg/ml TBZ for five hours, fixed with paraformaldehyde and stained with Calcofluor. The septation index is the percentage of septated cells in the culture.