title: GSM1833967: Day 6, Animal 74; Bos taurus; RNA-Seq
library_construction_protocol: Whole blood was collected at DAY 0–7, 10, 14 and 21 and immediately mixed at a 3:5 (blood:buffer) ratio with RNALater (Invitrogen). Samples were stored at –20°C until processing. To purify RNA, frozen samples were thawed at 37°C, centrifuged at 2000xG, and supernatant discarded. Remaining cell pellets were subjected to two rounds of treatment with Red Blood Cell Lysing Buffer per manufacturer’s instructions (Sigma). RNA from resulting cell pellet was initially extracted by Trizol (Invitrogen) followed by treatment with RNeasy Kit with on-column DNase digestion (Qiagen). Purified RNA was quantified by spectrophotometry on a NanoDrop (Thermo, USA) and by electrophoresis with a BioAnalyzer 2100 (Agilent, USA). Only samples with an RNA Integrity Number (RIN) greater than 8.0 were used for RNASeq analysis. All RNASeq sequencing was performed at the Texas A&M Genetics and Bioinformatics Center following manufacturer’s instructions. cDNA was generated from RNA samples of sufficient quantity and quality with Ovation RNA-Seq System (NuGen, USA), sheared by focused ultrasound (Covartis, USA) and made into barcoded (multiplexed) libraries with Encore Rapid Library System (NuGen, USA).  Samples were quantified, diluted, pooled re-analyzed with a Bioanalyzer and loaded onto a Genome Analyzer II High-throughput sequencer (Illumina, USA).