GSM6481124
Total RNA was isolated from freshly dissected retina lysates using the RNeasy Mini Kit (QIAGEN). The mRNA was then purified from 1μg of total RNA using the Dynabeads® mRNA DIRECT™ Micro Kit (Thermo Fisher). Whole transcriptome libraries were prepared using the Ion Total RNA-seq Kit v2. The yield and size distribution of the amplified libraries were assessed with an Agilent Bioanalyzer using a DNA 1000 Kit. Sequencing was performed on an Ion Chef™ Instrument (Ion Torrent™, Thermo Fisher). Analysis was performed using the Torrent Suite software v5.4.0 (Thermo Fisher Scientific) and the RNASeqAnalysis plugins (Thermo Fisher Scientific). The RNASeqAnalysis plugins aligns reads on mouse reference genome (mm10) using STAR, and then unmapped reads are aligned using Bowtie2 and merged together. Gene level expression is calculated using HTSeq and Picard to report raw read counts and FPKM (fragment per kilobase of transcript per million).