Mating Top2G-GFP and Spo11-FZZ expressing cells (107 cells) at meiotic prophase (3 h) and the post-meiotic stage (6 h) were collected by centrifugation following pretreatment with 0.5 mM PMSF (Roche Diagnostics, Indianapolis, IN) for 30 min at 30°C. Cells were resuspended in 1 ml homogenization buffer (150 mM NaCl, 1% Triton X-100 [Sigma-Aldrich], 2 mM PMSF, and Complete Protease Inhibitor Cocktail [Roche Diagnostics]), homogenized by gentle pipetting on ice, and clarified by centrifugation at 10,000 g for 15 min. The resulting lysate was incubated with 25 μl GFP-Trap magnetic agarose beads (ChromoTek, Planegg-Martinsried, Germany) or 1 μg anti-FLAG antibody bound to 25 μl Protein G magnetic agarose beads (GE Healthcare) for 1 h at 4°C. After three washes with homogenization buffer, the beads were incubated with 30 μl SDS sample buffer at 98°C for 2 min to elute bound proteins. Samples (15 μl) of eluted proteins were separated by SDS-PAGE. WCL: whole cell lysate, IP, immunoprecipitation fraction.