GSM9012089: U-CTRL, sow 20; Sus scrofa; RNA-Seq
Total RNA was extracted from frozen uterine tissue using Trizol reagent (Invitrogen, Waltham, 115 MA, USA) according to the Novogen protocol. RNA quantity was determined using the Qubit fluorometer (Life Technologies), RNA quality with the Agilent Bioanalyzer 2100 (Agilent Technologies), and RNA purity with NanoDrop (Thermo Scientific). RNA libraries were prepared for all 20 samples. First, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Then, libraries were prepared using Novogene NGS RNA Library Prep Set (PT042). After fragmentation, the first cDNA strand was synthesized using random hexamer primers. The second cDNA strand was synthesized using either dUTP for directional library or dTTP for non-directional library. Prepared libraries were quantified using the Qubit fluorometer and real-time PCR, and size distribution was assessed by the Agilent Bioanalyzer 2100, using the Agilent DNA 1000 kit. Quantified libraries were pooled and sequenced by Illumina Novaseq6000 S4 sequencer generating 150 bp paired end reads