AbstractIn the meiotic prophase, programmed SPO11‐linked DNA double‐strand breaks (DSBs) are repaired by homologous recombination (HR). The MRE11‐RAD50‐NBS1 (MRN) complex is essential for initiating DNA end resection, the first step of HR. However, residual DNA end resection still occurs in Nbs1 knockout (KO) spermatocytes for unknown reasons. Here, we show that DNA end resection is completely abolished in Mre11 KO spermatocytes. In addition, Mre11 KO, but not Nbs1 KO, undifferentiated spermatogonia are rapidly exhausted due to DSB accumulation, proliferation defects, and elevated apoptosis. Cellular studies reveal that a small amount of MRE11 retained in the nucleus of Nbs1 KO cells likely underlies the differences between Mre11 and Nbs1 KO cells. Taken together, our study not only demonstrates an irreplaceable role of the MRE11 in DNA end resection at SPO11‐linked DSBs but also unveils a unique function of MRE11 in maintaining the long‐term viability of undifferentiated spermatogonia.