GSM7502137: PDX, baseline UCLHN04 biol rep3; Homo sapiens; RNA-Seq
RNA extraction with phenol/chloroform and purification on RNeasy mini columns (Qiagen), according to manufacturer's instructions. Libraries are prepared with NEBNext UltraII Directional RNA Library Prep Kit for Illumina protocol according supplier recommendations. Briefly the key stages of this protocol are successively, the purification of PolyA containing mRNA molecules using poly-T oligo attached magnetic beads from 1μg total RNA (with the Magnetic mRNA Isolation Kit from NEB), a fragmentation using divalent cations under elevated temperature to obtain approximately 300bp pieces, double strand cDNA synthesis and finally Illumina adapters ligation and cDNA library amplification by PCR for sequencing. Sequencing is then carried out on Paired End 100b reads of Illumina NovaSeq.Image analysis and base calling is performed using Illumina Real Time Analysis (3.4.4) with default parameters.