GSM8164525: AB5E1 ALI7, patient 5; Homo sapiens; RNA-Seq
Total RNA was purified from frozen primary tumor cell pellets with RNeasy Plus Micro Kit (Qiagen, Hilden, Germany) according to the manufacturer's specifications. Libraries are prepared with NEBNext Ultra II 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.