RNAseq analysis of genes activated 90 minutes, or 7 days post irradiation or untreated cells with knockdown of NFKBIA

Goal of the study was to investigate genes regulated differentially after 90 minutes versus 7 days after irradiation. 7 day time point corresponds to cellular senescence. U2OS cells were irradiated with 20 Gy and harvested either 90 minutes or 7 days. In addition cells were left untreated (control) or had knockdown of NFKBIA (protein name Ikappa B alpha). Ikappa B alpha is the main inhibitor of the transcription factor NFkappa B. NFkappa B regulates transcription of many factors constituting the SASP (senescence associated secretory phenotype). Overall design: cell were grown in DMEM supplemented with 10 percent FCS and antibiotics. Cells (in quintuplicates) were irradiated with 20 Gy and harvested using Trizol reagent at time points indicated above (so that cell harvesting took place at the same time). RNA was analyzed for integrity and subseqently 500 ng was used from each sample for sequencing.