title: GSM3182004: Diploid_RNA_Seq_2; Mus musculus; ssRNA-seq
library_construction_protocol: Strand-specific nuclear RNA-seq library preparation Ten to 20 million unfixed ESCs (grown in 2i medium without feeder cells) were washed with cold PBS, resuspended in 0.5 ml of cold buffer RLN (50 mM Tris pH 7.5, 140 mM NaCl, 1.5 mM MgCl2, 1 mM DTT, 0.4% IGEPAL CA-630) and incubated for 5 min on ice. The samples were spun and the resultant pellets (nuclei) were washed with buffer RLN and subject to RNA isolation using RNeasy Mini Kit (Qiagen) and QIAshredder spin columns (Qiagen) according to the instructions from the manufacturer. The strand-specific RNA-seq libraries were prepared from 200 ng each of nuclear RNA samples using TruSeq Stranded Total RNA LT Sample Prep Kit (Illumina) according to manufacturer's instructions.