Chromatin immunoprecipitation of PRDM9 and H3K4me3

ChIP experiments were performed with the ChIP-IT High Sensitivity Kit (Active-motif, 53040). Briefly, testes from two or three synchronized mice (see above) were de-capsulated and fixed in complete tissue fixation solution for 10 min. After quenching the reaction, tissues were homogenized, and cell suspensions prepared by filtering samples through a 40 µm cell strainer. Cells were washed twice with ice-cold 1x PBS, and chromatin was extracted and immunoprecipitated according to the manufacturers’ instructions. 30–40 µg of chromatin was used per IP. The following antibodies (amount) were used: affinity purified anti-PRDM9 (4 µg), anti-H3K4me3 (4 µg).