Figure EV5. Conditional depletion of DDK before pachytene exit results in defective SC morphology.

• A
  Single colonies from the indicated strains were streaked on rich media with or without rapamycin. Since DBF4 and CDC7 are both essential genes, the observed growth defect in the presence of rapamycin indicates conditional inactivation of DDK.
• B
  The ndt80Δ DBF4‐FRB strain was induced to synchronously enter meiosis. At 6 h, either carrier (control) or rapamycin (rapamycin) was added. Cells were harvested 2 and 4 h after drug treatment, and chromosomes were spread as in Fig 4B. To quantify these results, nuclei were scored for the presence of fully established SC, as judged by Zip staining, and the presence of Red1 staining (for representative images, see control nuclei). Nuclei in white boxes are expanded to the right. White arrowheads depict polycomplexes, which were larger than usual because of prolonged cell cycle arrest. At least 100 nuclei were scored in the presence and absence of rapamycin at each time point. Data are represented as means from two experiments. Scale bar, 5 μm.
• C, D
  Strains were induced to synchronously enter meiosis. At 6 h, β‐estradiol was added to induce production of Cdc5. At the indicated time points, cells were harvested for detection of proteins as in Fig 1B (C). At the 7 and 8 h time points, cells were also harvested for immunoprecipitation with an anti‐Dbf4 antibody (D). 5× 4A denotes that fivefold more dbf4‐4A sample was loaded than DBF4. “−” and “+” indicate the exclusion and inclusion of antibody for IP, respectively, with the no antibody condition serving as a negative control. WCE, whole‐cell extract. Sol., soluble fraction.
• E, F
  Strains were induced to synchronously enter meiosis. At 6 h, β‐estradiol was added to induce production of Cdc5. Cells were harvested at the indicated time points, and proteins were detected as in Fig 1B.

Source data are available online for this figure.