GSM5158967: d149.Soy.50 [103265-001-093]; Homo sapiens; RNA-Seq
The RNA isolation was performed according to the manufacturer's protocol. RNA quality was assessed using fragment analysis using the DNF-471-500 standard sensitivity RNA kit from Agilent™. RNA quality number was determined with Prosize 2.0 (Data Analysis software 2.0.0.51). Strand-specific mRNA-seq libraries for the Illumina platform were generated using Illumina protocols and sequenced at BaseClear BV (Leiden, The Netherlands). Briefly: The NEBNext Ultra Directional RNA Library Prep Kit for Illumina was used to process the sample(s). The sample preparation was performed according to the protocol "NEBNext Ultra Directional RNA Library Prep Kit for Illumina" (NEB #E7420S/L). Briefly, mRNA was isolated from total RNA using the oligo-dT magnetic beads. After fragmentation of the mRNA, a cDNA synthesis was performed. This was used for ligation with the sequencing adapters and PCR amplification of the resulting product. The quality and yield after sample preparation was measured with the Fragment Analyzer (see appendices). The size of the resulting products was consistent with the expected size distribution (a broad peak between 300-500 bp). RNA-seq. Briefly: Clustering and DNA sequencing using the Illumina NextSeq 500 was performed according to manufacturer's protocols. A concentration of 1.6 pM of DNA was used. NextSeq control software 2.0.2 was used. Primary data analysis and results: Image analysis, base calling, and quality check was performed with the Illumina data analysis pipeline RTA v2.4.11 and Bcl2fastq v2.17. The sequencing experiments were performed at the following site(s): GenomeScan B.V., Plesmanlaan 1d, 2333 BZ, Leiden.