The screen is performed in 96well format with poly-L-Lysine coated plates for better cell attachment.

Day 1: HCT116 cell transfection

HCT116 wild-type and Mad2+/- cells were transfected using lipoRNAiMAXi transfection reagent, then cultured for 4 days.

Day 4: Ethanol fixation

Plates were fixed with 80% EthOH.

Staining and Readout

Cells were stained with pH2AX antibody and DAPI.

pH2AX nuclear intensity and total cell number per well were determined using the Acumen Explorer. Also, pH2AX nuclear spots were quantified using Cellomics Arrayscan.