GSM8851725: NPC cells, BIHi267-B, MILS patient, 10 µM Sildenafil  in 0.1% DMSO für 24 hours, replicate 2; Homo sapiens; RNA-Seq
RNA was purified using the NucleoSpin RNA Plus kit (Macherey-Nagel, Düren, Germany, #740984) and eluted in 30 µL RNase-free H2O. The RNA concentration was measured using the NanoDrop 2000 photometer and aliquots of 1-2 µg RNA were snap-frozen in liquid nitrogen and stored at -80°C. The construction of the eukaryotic strand-specific transcriptome library was performed according to the „DNBSEQ Eukaryotic Strand-specific Transcriptome Resequencing“ protocol of the  Beijing Genetics Institute (BGI). Briefly, 500 ng total RNA was enriched for polyA-carrying molecules using oligo-dT beads. The RNA was then fragmented and the first cDNA strand was synthesized using random primers. Second strand cDNA was synthesized using dUTP instead of dTTP and the synthesized cDNA was subjected to end repair and 3' adenylation. Adaptors were ligated to the ends of these 3' adenylated cDNA fragments, which were then amplified by PCR. After quality control, the library was circularized and amplified to produce DNA nanoballs (DNBs). The DNB library was then paired-end sequenced on the DNBSEQ-500 platform (DNBSEQ Technology, BGI, Beijing, China) with a read length of 150 bp. After removing of the sequencing adapters, the resulting FASTQ files were finally filtered for the desired length of 150 bp and for quality scores of Phred+33.