GSM5949780: AT/RT CHLA06 Control C; Homo sapiens; RNA-Seq
Total RNA was extracted from cell pellets using Qiagen (Germantown, MD) RNeasy mini kit and column DNase treatment. RNAseq libraries were prepared using Illumina (San Diego, CA) TruSeq stranded mRNA sample preparation kit following manufacturer's recommended procedure with minor modifications. Briefly, mRNA was purified from 15ng of total RNA using oligo dT RNA purification magnetic beads. The purified mRNA was then fragmented at 94°C for 8 minutes, primed with random primers, and converted to double strand cDNA. The resulting cDNA was end repaired, dA tailed, and ligated to TruSeq adapter (with unique dual index barcodes). Libraries were then PCR amplified using 15 cycles. The final libraries were quality checked, quantitated, and pooled in equal molar for sequencing on NovaSeq SP 100cycle flowcells for 2X50bp sequencing.