GSM5769089: OV90_EpCAMlow_3; Homo sapiens; RNA-Seq
RNA was harvested using Trizol reagent. RNA-Seq was performed at the Erasmus MC Center for Biomics (Rotterdam, The Netherlands) according to manufacturer's instructions (Illumina) and using the TruSeq RNA sample prep kit v2. Briefly, polyA containing mRNA molecules were purified using poly-T oligo attached magnetic beads. Following purification, the mRNA was fragmented into ~200 bp fragments using divalent cations under elevated temperature. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This was followed by second strand synthesis using DNA polymerase I and RNaseH treatment. These cDNA fragments were end repaired, a single A base was added and Illumina adapters were ligated. The products were purified and size selected on gel and enriched by PCR. The PCR products were purified by Qiaquick PCR purification and used for cluster generation according to the Illumina cluster generation protocols (www.illumina.com).