GSM7093469: Ctrl_F1_blastocyst_RNA_rep2-7; Mus musculus; RNA-Seq
Acidic Tyrode's solution (Sigma) containing 0.5% PVP and 50 mM NaCl was used to remove the zona pellucida (ZP) from embryos before sample collection. Preparation of total RNA-seq libraries of pooled 2-cell and single XY blastocyst embryos was performed using SMART-Seq Stranded Kit (Clontech), according to the manufacturers' instructions. In short, pooled 30 each of ZP-free 2-cells per replicate and each single blastocyst in ICSI-derived Ctrl-F1 and Scml2-KO-F1 embryos, were lysed in 1x Lysis Buffer containing RNase inhibitor (0.2 IU/µl, from SMART-Seq Stranded Kit, Clontech), directly. RNAs were randomly sheared by heating at 85℃ for 6 min and supplied to reverse transcription with random hexamers and PCR amplification. Ribosomal fragments were depleted from each cDNA sample with scZapR and scR-Probes. Indexed total RNA-seq libraries were enriched by second PCR amplification.