GSM8682992: RNA_Liver_Y1_Rep2; Mus musculus; RNA-Seq
Mouse liver, kidney or brain were dissected from the euthanized mice under the Northwestern University protocol. Immediately after collection of these organs, they were snap frozen in liquid nitrogen for future use. For RNA extraction, the tissue was minced thoroughly in 1X PBS and then divided into 4 independent fractions. RNA was extracted from each fraction according to Qiagen RNeasy (Qiagen, 74104) protocol. Briefly, 1ml RLT buffer (containing 2-Mercaptoethanol ) was added to each minced tissue fraction, and then suspension was further homogenized using an electric homogenizer. Next, an equal volume of 100% molecular grade Ethanol was added to the mix and the entire volume was applied to RNeasy RNA extraction columns. After subsequent on-column DNase I treatment using Qiagen RNase-Free DNase Set (Qiagen, 79254), the columns were washed with the RNeasy kit buffers and the RNA was eluted in 50ul of the provided elution buffer. Next, RNA was subjected to ribosomal RNA depletion using the Illumina Ribo-Zero Plus rRNA depletion Kit (Illumina, 20036696), and libraries were synthesized using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs, E7760L). RNA-seq libraries were sequenced on an Illumina NovaSeq 6000 and NextSeq 2000 instrument using paired-end 50 cycle reads.