Porcine and bovine oocytes were obtained from the slaughterhouse material as described previously (Blaha et al, 2015 (link); Kinterova et al, 2019 (link)). For reporter injection, a mixture of in vitro‐transcribed firefly and nanoluciferase (NanoLuc) RNA in the ratio of 100,000:10,000 was injected per oocyte with FemtoJet microinjector (Eppendorf). For antisense oligonucleotide inhibitor injection, commercially obtained hsa‐let‐7a‐5p or hsa‐miR‐205‐5p miRCURY LNA miRNA inhibitor (Qiagen, cat# YI04101776 and YI04101508, respectively) was diluted in water and microinjected (~5 × 106 molecules per oocytes).
Injected bovine oocytes were cultured in MPM media (prepared in‐house (Kinterova et al, 2019 (link))) containing 1 mM dbcAMP (Sigma) without a paraffin overlay in a humidified atmosphere at 39°C with 5% CO2 for 20 h. Porcine oocytes were cultured in M‐199 medium (Gibco) supplemented with 1 mM dbcAMP, 0.91 mM sodium pyruvate, 0.57 mM cysteine, 5.5 mM HEPES, antibiotics, and 5% fetal calf serum (Sigma). Injected oocytes were incubated at 38.5°C in a humidified atmosphere of 5% CO2 for 20 h.
Injected bovine oocytes were cultured in MPM media (prepared in‐house (Kinterova et al, 2019 (link))) containing 1 mM dbcAMP (Sigma) without a paraffin overlay in a humidified atmosphere at 39°C with 5% CO2 for 20 h. Porcine oocytes were cultured in M‐199 medium (Gibco) supplemented with 1 mM dbcAMP, 0.91 mM sodium pyruvate, 0.57 mM cysteine, 5.5 mM HEPES, antibiotics, and 5% fetal calf serum (Sigma). Injected oocytes were incubated at 38.5°C in a humidified atmosphere of 5% CO2 for 20 h.
