title: GSM6124523: Emricasan - Inadequate Hepatic Function Liver - 3 hours perfusion - replicate 3; Homo sapiens; RNA-Seq
library_construction_protocol: Tissues samples from livers taken immediately prior to perfusion and after 3 and 6 hours of perfusion were used for bulk transcriptome sequencing. Core needle biopsies from the right liver lobe were collected in RNAlater solution (Sigma-MilleporeMillipore, Waltham, MA, USA) and stored overnight at 4 Celsius. Tissue samples were then removed from solution and stored at -80 Celsius. Total RNA was isolated using the RNeasy Mini Kit (Qiagen, Germantown, MD, USA) according to manufacturer guidelines. RNA sequencing libraries were prepared using the NEBNext Ultra RNA Library Prep Kit for Illumina following manufacturer's instructions (NEB, Ipswich, MA, USA). Briefly, mRNAs were first enriched with Oligo(dT) beads.  Enriched mRNAs were fragmented for 15 minutes at 94 °C. First strand and second strand cDNAs were subsequently synthesized. cDNA fragments were end repaired and adenylated at 3'ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by limited-cycle PCR. The sequencing libraries were validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA).