Illumina HiSeq 4000 paired end sequencing; Tissue-autonomous regulation of adipose expansion by core clock protein REVERBα
Gonadal white adipose tissue was collected from adult male ReverbαFlox2-6AdipoCre+/-, Reverbα-/- global knockout (KO) or Reverbα wildtype mice (littermates of global knockouts) at ZT8, and flash frozen (n=6-8). Frozen tissue was homogenised in TRIzol Reagent (Invitrogen) using Lysing Matrix D tubes (MP Biomedicals) and total RNA extracted according to the manufacturer's TRIzol protocol as far as the isopropanol phase, at which point samples were transferred to columns from the ReliaPrep RNA Miniprep Tissue Kit (Promega) to ensure high quality RNA.  The column was then washed, DNAse treated and RNA eluted as per Promega's protocol. Biological replicates were taken forward individually. Total RNA was submitted to the University of Manchester Genomic Technologies Core Facility (GTCF). Quality and integrity of the RNA samples were assessed using a 2200 TapeStation (Agilent Technologies) and then libraries generated using the TruSeq® Stranded mRNA assay (Illumina) according to the manufacturer's protocol.