Measurement of DNA Repair Pathways

HR and NHEJ assays were performed according to previous reports [38 (link)]. In brief, NHEJ reporter plasmid pimEJ5GFP (Addgene, #44026, USA) or HR reporter plasmid DR-GFP (Addgene, #26475) was transfected in HEK 293T cells. To check the effect of DJ-1 depletion on NHEJ and HR efficiency, cells were transfected with either DJ-1 siRNA or control. The pCBASceI plasmid (Addgene, #26477) was transfected into both control and DJ-1 siRNA-treated cells at 48 h post-transfection. After 24 h, cells were harvested and subjected to flow cytometry (Beckman CytoFLEX). To investigate the effect of DJ-1 overexpression on NHEJ and HR efficiency, cells were transfected with empty vector/ Flag-DJ-1 and pCBASceI plasmids. Cells were collected after 48 h, and the GFP signal was evaluated by flow cytometry.