Figure 5.

DSBs persist late into prophase in top2-1 mutants. (A) DNA content of dmc1Δ and dmc1Δ top2-1 cells as determined by flow cytometry. Samples were taken at the indicated time point. (B) Southern blot analysis of DSBs throughout a meiotic time course across chromosome VIII in wild-type and top2-1 cells. Cultures were shifted to the restrictive temperature (34°) 1h after meiotic induction. (C) Background signal was subtracted from DSB signal and each experiment was normalized to the maximal signal of the control. Average and SD for three experiments are plotted. (D–F) Time course analysis of wild-type and top2-1 cells. The analysis methods are the same as in A–C. (G) Spindle pole separation as measured by anti-tubulin immunofluorescence of wild-type (black), top2-1 (blue), spo11-Y135F (red), and top2-1 spo11-Y135F (purple) cells (n = 200 for each data point). (H) Representative images of immunofluorescence staining for Rad51 (green) and Zip1 (red) on chromosome spreads of wild type and the top2-1 mutant at the restrictive temperature (34°) 4–5 hr after meiotic induction. Bar, 5 μm.