GSM5090263: SK-N-BE(2) xenograft - 4SC-205_rep2; Homo sapiens; RNA-Seq
Tumors were excised and total RNA was isolated using the miRNeasy mini kit (Qiagen). Qubit® RNA HS Assay (Life Technologies) and RNA 6000 Nano Assay on a Bioanalyzer 2100 (Agilent) were used to determine the total RNA quality and quantity. The RNASeq libraries were prepared following the TruSeq® Stranded mRNA LT Sample Prep Kit protocol (Illumina). Total RNA (500ng) was enriched for the polyA mRNA fraction and fragmented by divalent metal cations at high temperature. In order to achieve the directionality, the second strand cDNA synthesis was performed in the presence of dUTP. The blunt-ended double stranded cDNA was 3´adenylated and Illumina platform compatible adaptors with unique dual indexes and unique molecular identifiers (Integrated DNA Technologies) were ligated. The ligation product was enriched with 15 PCR cycles and the final library was validated on an Agilent 2100 Bioanalyzer with the DNA 7500 assay (Agilent).The libraries were sequenced on NovaSeq 6000 (Illumina) with a read length of 2x51bp.