title: GSM8333216: Liver, Placebo, d0, patient 116; Homo sapiens; RNA-Seq
library_construction_protocol: Total RNA from transjugular liver biopsies was extracted using QIAzol lysis reagent protocol (Qiagen). RNA samples were treated with Turbo DNA-free Kit (Thermo Fisher) to remove genomic DNA following manufacturer's guidelines. Where possible 10ng of each total RNA sample was fragmented to a size appropriate for sequencing on an Illumina platform and first-strand cDNA was generated using the SMARTer Stranded Total RNA-Seq Kit v2 – Pico Input Mammalian kit (Clontech Laboratories, Inc. #634411). Samples were grouped for library prep based on the amount of fragmentation required, as recommended in the protocol. [n=8 samples were subject to 4 minutes fragmentation time; n=15 samples were subject to 3 minutes of fragmentation; remaining samples required no fragmentation.] Illumina-compatible adapters and indexes were then added via 5 cycles of PCR. AMPure XP beads (Beckman Coulter, #A63881) were then used to purify the cDNA library. Depletion of ribosomal cDNA was performed using ZapR v2 and R-probes v2 specific to mammalian ribosomal RNA and human mitochondrial rRNA. Uncleaved fragments were enriched by 13 cycles of PCR before a final library purification using AMPure XP beads.