Analysis of gene expression in Idasanutlin-arrested SIN3A and SIN3B depleated cells
mRNA expression analysis of arrested HCT116 cells (wild-type or SIN3B-/-) transfected with either non-targeting control siRNA or one of three SIN3A siRNAs and treated with Idasanulin to activate p53. Overall design: HCT116 cells were cultivated in 6-well plates and transfected with 20 nM SIN3A siRNAs (Horizon Discovery, #MQ-012990-00-0002; siRNAs #1, #2, and #4 for three biological replicates) or a non-targeting control siRNA (siGENOME Non-Targeting siRNA 5, Horizon Discovery; three biological replicates) and 5 µl Lipofectamine RNAiMAX (Invitrogen, #13778075) in a total volume of 2 ml in antibiotics free 10% DMEM. 24 h after transfection, cells were treated with either DMSO or 5 µM Idasanutlin (R&D Systems, #12-35-22-07) for an additional 24 h before cells were harvested RNA extraction. Total RNA was isolated with the Direct-zol RNA MiniPrep Kit (Zymo Research, #R2053).