alias: GSM3530840
title: GSM3530840: WGBS Foxa1L/L;2L/L  2; Mus musculus; Bisulfite-Seq
library_construction_protocol: Mice were anesthetized, sacrificed, and subjected to liver perfusion. Whole liver was then digested by collagenase (Roche) and hepatocytes removed and isolated by means of a percoll gradient (18). This yielded a population of relatively pure (>95%) hepatocytes. DNA was isolated from mouse hepatocytes or from snap-frozen mouse tissues and incubated in lysis buffer (25 mM Tris-HCl (pH 8), 2 mM ethylenediaminetetraacetic acid, 0.2% sodium dodecyl sulfate, 200 mM NaCl) supplemented with 300 μg/mL proteinase K (Roche) followed by phenol:chloroform extraction and ethanol precipitation. WGBS libraries were prepared using the Ovation® Ultralow Methyl-Seq Library System (NuGEN Technologies Inc.) as per the manufacturer's instructions. One hundred and fifty nanograms of genomic DNA was sheared with the M220 Focused-ultrasonicator™(Covaris®) using the 200 bp snap-cap protocol and subsequently purified with AMPure XP beads. After end repair, adaptor ligation, and final repair, DNA was bisulfite converted with the EpiTect Fast DNA Bisulfite Kit (Qiagen GmbH) following the cycling conditions specified in the Ovation® Ultralow Methyl-Seq Library System protocol. Libraries were then amplified, purified, pooled, and sequenced on the Illumina HiSeq with 100 bp single-end sequencing. Sequencing was conducted by the University of Pennsylvania Next-Generation Sequencing Core (Philadelphia, PA, USA) at a depth that exceeds the 20-fold commonly achieved in WGBS databases.
identifiers.namespace: GEO
identifiers.id: GSM3530840