GSM2546925: C5; Mus musculus; RNA-Seq
Epididymal adipose tissue was rapidly excised, flash frozen in liquid nitrogen and crushed into a homeogeneous powder prior to isolation of RNA.  RNA was isolated using TRI Reagent with modifications to remove DNA using Qiagen RNAeasy columns and RNAase-free DNaseI. Quality and quantity of RNA was determined using an Agilent 2100 Bioanalyzer. RNA will be transcribed into cDNA via priming at the poly A+ tail. Double-stranded cDNA will be digested with NlaIII restriction enzyme and ligated to adaptors containing an EcoP15L restriction site. Digestion with EcoP15L, ligation of "bar code" oligonucleotides that specifically label each sample, and subsequent PCR amplification will yield a library for sequencing where each gene will be represented by a 3'-expression tag.