Wild type c57bl 6 j female mice

Oocyte preparation Wild-type C57BL/6 J female mice of 5–8 weeks (Janvier Labs, Le Genest-Saint-Isle, France) were superovulated with 5 IU of pregnant mare serum gonadotropin (PMSG) and then 48 h later with 5 IU human chorionic gonadotropin (hCG) (Intervet, Beaucouzé, France). The next day, about 13 h after hCG injection, animals were sacrificed by cervical dislocation. Cumulus oocyte complexes were collected by tearing the ampulla wall of the oviduct, placed in FertiCult medium (FertiPro N.V, Belgium) supplemented with 3% BSA (Sigma–Aldrich), and maintained at 37 °C under 5% CO₂ in air under FertiCult Mineral Oil (FertiPro N.V, Belgium). When experiments were performed with zona-free oocytes, cumulus cells were first removed by a brief exposure to hyaluronidase IV-S (1 mg/ml, Sigma–Aldrich). Then, the zona pellucida was dissolved with acidic Tyrode’s solution (pH 2.5, Sigma–Aldrich) under visual monitoring. Zona-free eggs were rapidly washed in a FertiCult medium, BSA 3% and kept at 37 °C under 5% CO₂ atmosphere for 2 to 3 h to recover their fertilization ability.
Sperm preparation Mouse spermatozoa were obtained from the cauda epididymis of WT or Tex44-KO male mice (8–14 weeks old) and capacitated at 37 °C under 5% CO₂ for 90 min in a 500 μl drop of FertiCult medium supplemented with 3% BSA, under FertiCult Mineral Oil.

Sexually mature (8 to 14 weeks) Tex44-KO homozygous and wild-type littermates were mated with C57BL/6 J females of 7–8 weeks. The numbers of pups and litters were recorded after 3 weeks of gestation whose beginning was attested by the presence of a vaginal plug after the overnight mating.
Wild-type C57BL/6 J female mice of 5–8 weeks (Janvier Labs, Le Genest-Saint-Isle, France) were injected with 5 IU of pregnant mare serum gonadotropin (PMSG). After 48 h, they were superovulated with 5 IU human chorionic gonadotropin (hCG) (Intervet, Beaucouzé, France) and then mated overnight with sexually mature WT or Tex44-KO males in order to evaluate their capacity to fertilize in vivo. The next day, females showing a vaginal plug were sacrificed by cervical dislocation and their oocytes retrieved. Oocytes were cleared of cumulus cells and directly mounted in Vectashield/DAPI (Vector laboratories, Burlingame, CA, USA) for observation under UV light (Nikon Eclipse E600 microscope) in a blinded manner. Oocytes showing at least one fluorescent decondensed sperm head within their cytoplasm were considered fertilized. According to this, the fertilization rate (FR) was evaluated.