title: Illumina NovaSeq 6000 paired end sequencing; RNA-seq of livers from mice treated for eight weeks with the glucagon receptor antibody, REGN1193, compared to mice treated with a control antibody, REGN1945, (control group)
design: RNA-seq of livers from mice treated for eight weeks with the glucagon receptor antibody, REGN1193, compared to mice treated with a control antibody, REGN1945, (control group)
library_name: Sample 67_p
library_construction_protocol: The mice were fasted overnight (11 hours) prior to liver isolation, and livers were kept at -80°C until analysis.  For eight weeks, female C57BL/6JRj mice (seven weeks old at the start of treatment) received, by subcutaneous injections, either a glucagon receptor antibody (GCGR Ab, REGN1193, 10 mg/kg body weight; Regeneron; IC50: 3.5 pM (Okamoto et al., 2015)) or a control antibody (Ctl Ab, REGN1945, 10 mg/kg body weight; Regeneron). The antibodies were administered once weekly (Wednesdays at 8 AM). The last doses of GCGR Ab and Ctl Ab were given one week prior to termination. Raw sequencing data were processed using bcl2fastq software version 2.20.0 (Illumina). Total DNA/RNA was purified by an AllPrep DNA/RNA Minikit (80204, QIAgen) according to manufacturer's instructions and quality tested using a 2100 Bioanalyzer instrument (Agilent Genomics). The extract was DNase treated with RNeasy Minikit (74106, QIAgen) and RNase-Free DNase set (79254, QIAgen) according to manufacturer's protocol. All the above-mentioned processes were performed on a QIACube machine (QIAgen).  Library preparation was done using TruSeq Stranded Total RNA Library Prep Gold (20020599, Illumina) and IDT for Illumina-TruSeq RNA UD Indexes (20022371, Illumina) according to the manufacturer's instructions. Some samples (input <100 ng) were prepared with NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (NEB-E7760S, New England Biolabs), NEBNext rRNA Depletion Kit 24 with beads (NEB-E7765, New England Biolabs), and NEBNext Multiplex Oligos for Illumina (E7600S, New England Biolabs).