BT549 STAT1 KO Reversine #2_p
For Reversine/DMSO treatment: BT549 cells (with according KOs) were synchronized with double thymidine block. Seven hours after thymidine release, synchronized BT549 cells were treated with DMSO or 250 nM Reversine. After 48 hours, cells were harvested using Tryple. Pelleted cells were washed with 1xDPBS before being stored as dry pellets at -20 degree Celsius. For IFN gamma treatment: BT549 cells were treated with 50 ng IFN gamma for 2 hours. After 2 hours incubation, cells were harvested and washed with 1XDPBS before being stored as dry pellet at -20 degree Celsius. For whole transcriptome analysis, RNA was isolated using the RNeasy mini kit (Qiagen) in accordance to manufacturer's protocol and RNA quality was examined using the Agilent 2100 Bioanalyzer (Agilent Technologies). 250-500 ng of total RNA was used as input for mRNA enrichment using NEXTflex Poly(A) Beads (Bio Scientific) followed by library preparation using a NEXTflex Rapid Directional qRNA-Seq Kit (Bio Scientific).