Immunostaining of Meiotic Proteins

For primary antibodies, we used previously generated guinea pig anti-HORMAD1 (1:100, [41 (link)]), anti-SYCP3 (1:200, gift from R. Benavente, [80 (link)]) and rabbit polyclonal anti-RAD51 (1:1000, [81 (link)]), anti-SYCP1 (1:2500, [82 (link)]) and mouse monoclonal anti-DMC1 (1:1000, Abcam, ab11054), anti-SYCP3(1:200, Abcam, ab97672), For secondary antibodies, we used goat anti-guinea pig IgG Alexa 647 (1:500, Abcam, ab150187), goat anti-rabbit IgG Alexa 647 (1:250, Invitrogen, A21245), goat anti-rabbit IgG CF568 (1:500, Sigma Aldrich, SAB4600310), goat anti-mouse IgG CF568 (1:500, Biotium, 20109) and goat anti-mouse IgG Atto 488 (1:250, Rockland, 610-152-121S). For STED microscopy we used a goat-anti mouse IgG Alexa 555 (1:500, Invitrogen, A-21422). Control immunostainings in Dmc1^-/- and Spo11^-/- spermatocytes, which were imaged using a Zeiss Confocal Laser Scanning Microscope 700 with a 63x objective immersed in oil (images were taken with the same intensity), confirmed loss of DMC1 signal, and loss of signal of both recombinases respectively (S9 Fig).