GSM4257114: Ctrl  RNA-seq rep2; Mus musculus; RNA-Seq
RNA was extacted using Trizol following manufacturer's protocol. For cDNA synthesis, custom oligo-dT primers were used with a barcode and adapter-linker sequence (CCTACACGACGCTCTTCCGATCT—XXXXXXXX-T15). After first strand synthesis, samples were pooled together based on Actb qPCR values, and RNA-DNA hybrids degraded using consecutive acid-alkali treatment. A second sequencing linker (AGATCGGAAGAGCACACGTCTG) was ligated using T4 ligase (NEB) followed by SPRI clean-up. The mixture was then PCR enriched for 12 cycles and SPRI purified to yield final strand specific RNA-seq libraries as previously described (Jha et al., 2015) Library strategy: Libraries were sequenced using a HiSeq 2500 (Illumina) using 40 bp by 10 bp single-end sequencing.