GSM3271941: Liver-RE ZT08 Replicate 2; Mus musculus; RNA-Seq
Total RNA was extracted from liver with TRIzol (Invitrogen), followed by precipitation with isopropanol and ethanol. Library construction was performed according to the Illumina TruSeq Total RNA stranded protocol.  The input quantity for total RNA was 1ug and rRNA was depleted  using ribo-zero rRNA gold removal kit (human/mouse/rat). The rRNA depleted RNA was chemically fragmented for three minutes.  First strand synthesis used random primers and reverse transcriptase to make cDNA.  After second strand synthesis the ds cDNA was cleaned using AMPure XP beads and the cDNA was end repaired and then the 3' ends were adenylated.  Illumina barcoded adapters were ligated on the ends and the adapter ligated fragments were enriched by nine cycles of PCR. The resulting libraries were validated by qPCR and sized by Agilent Bioanalyzer DNA high sensitivity chip.  The concentrations for the libraries were normalized and then multiplexed together.  The concentration for denaturation was 2nM and the final concentration for clustering was 200pM.  The multiplexed libraries were sequenced by the Genomics High-Throughput Facility at the University of California – Irvine on six lanes using single end 100 cycles chemistry for the HiSeq 4000.   The version of HiSeq control software was HCS 3.4.0.38 with real time analysis software, RTA 2.7.7.