NextSeq 500 sequencing; RNA-seq of human CLB-GA and NGP neuroblastoma cell lines with SOX11 knockdown as compared to control
RNA-seq of human CLB-GA and NGP neuroblastoma cell lines with SOX11 knockdown as compared to control
Cell lines were harvested, centrifuged and stored at -80°C. Cells were grown in RPMI1640 medium supplemented with 10% foetal bovine serum (FBS), 2 mM L-Glutamine and 100 IU/ml penicillin/streptavidin at 37°C in a 5% CO2 humid atmosphere Cells were seeded in 6-well tissue culture plates 24 hours prior to transfection. 100nM of siRNA non-targeting control (siRNA NTC, D-001810-10-05) or siRNA SOX11 (L-017377-01-0005, Dharmacon) were transiently transfected using DharmaFect 2 (Thermo Fisher Scientific) according to the manufacturer's guidelines. For nucleofection, cells were nucleofected with 100 nM of the above described siRNA NTC and siRNA SOX11 using the Neon Transfection System (Thermo Fisher Scientific) and subsequently seeded in 6-well or T25 tissue culture plates. Total RNA was extracted using miRNeasy kit (Qiagen) according to manufacturer's instructions, including on-column DNase treatment, and concentration was determined with the Nanodrop (Thermo Scientific). CLB-GA samples were prepared using the TruSeq Stranded mRNA Sample Prep Kit from Illumina. And NGP samples were prepared using the QuantSeq 3' mRNA-Seq Library Prep Kit from Illumina.