title: GSM1432231: RNA-seq of PolyA Minus RNA from  Mouse ES cells  with Floxed ATRX Allele; Mus musculus; RNA-Seq
library_construction_protocol: RNA was isolated with Tri-Reagent (Sigma). The RNA quality was assessed using Agilent Bioanalyser. Only RNA with overall RIN score >9 was used for further procedures. Subsequently, RNA was treated with DNaseI (DNA-free, Ambion) according to the manufacturer’s protocol kit according to the protocol. Good quality and DNA-free RNA was subjected to polyA fractionation using PolyA selection kit (Promega) using the standard kit protocol.  The RNA fraction was precipitated overnight and resuspended with water. The quality of obtained RNA samples was assessed using PicoChip (Agilent). A Paired sequencing library was generated using the standard Illumina prtocol. Prior to library prep, the polyA- samples were treated with EpiCentre's Ribo-Zero™ Magnetic Kit (Human/Mouse/Rat). All libraries were prepped with the following kit. NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®. The protocol was modified to make use of our own custom 8bp indexes (Lamble et al. 2013). The prepared libraries then sequenced on a HiSeq2500 in rapid mode (no version number as original chemistry), 50bp paired end.