Chromosome Spread Preparation and Imaging

Spermatocyte spreads were prepared as we previously described^{65 (link),66 (link)}. In brief, the seminiferous tubules were treated with hypotonic extraction buffer for 30 min, and then germ cells were squeezed out in one drop of 100 mM sucrose solution and spread on slides with 1% PFA containing 0.15% Triton X-100. The slides were placed in a humidified chamber for at least 2 h and air-dried. Oocyte spreads were prepared from ovaries of 16.5–18.5 dpc female mice as previously reported^{9 (link),67} . Immunofluorescence staining was carried out as we previously described^{52 (link)}. The primary and secondary antibodies used and their dilutions are shown in Supplementary Table S3. Conventional fluorescence images were captured with an Olympus BX53 microscope (Olympus, Tokyo, Japan) with a scientific complementary metal-oxide-semiconductor camera (Prime BSI, Teledyne Photometrics Inc., USA) and processed with the Olympus cellSens software. Super-resolution images were captured using structured illumination microscopy (Nikon, N-SIM) equipped with a 100× oil-immersion objective lens (SR Apo TIRF 100×, NA 1.49) and a CCD camera (Andor, DU-897, X-11459), and images were processed using the NIS-Elements software.