title: GSM4105607: Control Endpoint replicate 1; mouse gut metagenome; RNA-Seq
library_construction_protocol: Mice were euthanized, the cecum dissected and contents collected under sterile conditions Cecal contents at 6 weeks of age and at study endpoint. Samples were immediately frozen and stored at -80oC until analysis. DNA from 50 mg of cecal contents was isolated from each mouse using DNeasy PowerSoil Kit (Qiagen). DNA sequencing was performed by the Centre for the Analysis of Genome Evolution and Function (Toronto). The V4 hypervariable region of the 16s rRNA gene was amplified using a universal forward sequencing primer and a uniquely barcoded reverse sequencing primer to allow for multiplexing.22 All amplification reactions were done in triplicate, checked on a 1% agarose TBE gel and then pooled. The final library was purified using 0.8X magnetic Ampure XP beads, selecting for the bacterial V4 amplified band. The purified library was quantified and loaded on to the Illumina MiSeq for sequencing, according to manufacturer's instructions (Illumina, San Diego, CA). Sequencing was performed using the V2 (150bp x 2) chemistry.