Detailed Western Blotting Protocol Using Phos-tag

Western blotting was carried out as described^{23 (link)}. Phos-tag SDS-PAGE gels were made from 7% acrylamide (37.5:1 crosslinking ratio), 70 μM Phos-tag reagent (Wako Pure Chemical Industries) and 140 μM MnCl₂, and were processed according to the manufacturer’s manual. Proteins were detected using the following primary antibodies: mouse anti-GFP (1:1000, Roche 11814460001), mouse anti-PLK1 (1:2000, Santa Cruz sc-17783), mouse anti-histone H3 pSer10 (1:5000, Abcam 14955), mouse anti-α-tubulin (1:5000, Sigma-Aldrich T9026), rabbit anti-phospho-(Ser) CDK substrate (1:1000, Cell Signaling 2324), rabbit anti-histone H3 (1:2000, Abcam ab1791), mouse anti-FLAG HRP (1:1000, Sigma-Aldrich A8592), mouse anti-MUS81 (1:1000, Santa Cruz sc-47692), mouse anti-BLM (1:500, Santa Cruz sc-70426), rabbit anti-KAP-1 pSer842 (1:1000, Abcam ab70369), rabbit anti-CHK1 pSer345 (1:1000, Cell Signaling 2341), mouse anti-histone H2A.X pSer139 (1:1000, Millipore 05-636-1), rabbit anti-GEN1 (1:100, raised against GEN1^890-908)¹⁹ . Uncropped blots are show in Supplementary Fig. 5.