GSM5819080: NSC_T13_2; Homo sapiens; RNA-Seq
Total RNA was extracted from frozen cell pellets using Qiagen RNeasy Mini kit following  manufacturer's instructions (Qiagen, Hilden, Germany).  RNA samples were quantified using Qubit 4.0 Fluorometer (Life Technologies,  Carlsbad, CA, USA) and RNA integrity was checked with RNA Kit on Agilent 5300  Fragment Analyzer (Agilent Technologies, Palo Alto, CA, USA) RNA sequencing libraries were prepared using the NEBNext Ultra RNA Library Prep Kit  for Illumina following manufacturer's instructions (NEB, Ipswich, MA, USA). Briefly,  mRNAs were first enriched with Oligo(dT) beads. Enriched mRNAs were fragmented  for 15 minutes at 94 °C. First strand and second strand cDNAs were subsequently  synthesized. cDNA fragments were end repaired and adenylated at 3'ends, and  universal adapters were ligated to cDNA fragments, followed by index addition and  library enrichment by limited-cycle PCR. Sequencing libraries were validated using NGS  Kit on the Agilent 5300 Fragment Analyzer (Agilent Technologies, Palo Alto, CA, USA),  and quantified by using Qubit 4.0 Fluorometer (Invitrogen, Carlsbad, CA).   The sequencing libraries were multiplexed and loaded on the flowcell on the Illumina  NovaSeq 6000 instrument according to manufacturer's instructions