NextSeq 500 sequencing; RNA-seq of CLB-GA neuroblastoma cell line upon TBX2 knockdown
RNA-seq of CLB-GA neuroblastoma cell line upon TBX2 knockdown
Cell lines were harvested, centrifuged and stored at -80°C. Cells were grown in RPMI1640 medium supplemented with 10% foetal bovine serum, 2 mM L-Glutamine and 100 IU/ml penicillin/streptavidin at 37°C in a 5% CO2 humid atmosphere Cells were transduced with concentrated virus (shTBX2_2, shTBX2_4 or NTC) and 24h after transduction cells were refreshed with medium. Puromycin selection was started 48h upon transduction. 6 days upon transduction, cells were harvested. 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). 250 ng of RNA isolated from cells was used as input for library preparation with the  TruSeq Stranded mRNA Sample Prep Kit from Illumina, followed by quality control on the bio-analyzer (Agilent) and qPCR quantificaiton of the libraries using the Illumina Kapa Library quantification kit (Ligtcycler 480 qPCR mix Kapa Code KK4854)
Experimental Factor: RNA interference
scrambled shRNA