GSM6742689: Liver, BMT, 3; Mus musculus; RNA-Seq
Tissue was harvested on day 7 post-transplant and bulk liver and isolated epithelial cells of the small and large intestine were suspended in TRIzol and homogenized with metal beads. Phase separation in homogenized tissue or in cells lysed in TRIzol was induced with 200 µL chloroform and RNA was extracted from the aqueous phase using the MagMAX mirVana Total RNA Isolation Kit (ThermoFisher catalog # A27828) on the KingFisher Flex Magnetic Particle Processor (Thermo Scientific catalog # 5400630) according to the manufacturer's protocol. Samples were eluted in 34 µL elution buffer. After RiboGreen quantification and quality control by Agilent BioAnalyzer, 500 ng of total RNA with RIN values of 7.2-10 underwent polyA selection and TruSeq library preparation according to instructions provided by Illumina (TruSeq Stranded mRNA LT Kit, catalog # RS-122-2102), with 8 cycles of PCR. Samples were barcoded and run on a NovaSeq 6000 in a PE100 run, using the NovaSeq 6000 S2 Reagent Kit (200 Cycles) (Illumina).  An average of 30 million paired reads was generated per sample. Ribosomal reads represented 0.2-0.9% of the total reads generated and the percent of mRNA bases averaged 93%.