GSM3664928: FB359; Homo sapiens; RNA-Seq
Cells at passages 3–4 were used for RNA extraction performed after 24 hr incubation in serum free medium. Total RNA was extracted from human SMC cultures with Trizol® Reagent (Life Technologies), purified by miRNeasy Mini kit® (Qiagen S.A.) according to the manufacturer's instructions. DNase digestion procedure was followed for each sample. Libraries were prepared from 1μg of total RNA using TruSeq Stranded mRNA kit protocol (Illumina®) according to the supplier's recommendations. Briefly, poly-A RNA was isolated using oligo-dT25 attached to magnetic beads, fragmented into ~300 SCP-long pieces using divalent cations under elevated temperature, used as template for double-stranded cDNA synthesis, Illumina adapters ligation and PCR-mediated cDNA library amplification. Sequencing was carried out on Illumina HiSeq4000,