Cell culture, cell lysis, immunoprecipitation, and western blotting

MiaPaCa-2 cells were obtained from the American Type Culture Collection (ATCC). HCT116 parental and FBXW7 isogenic cells were gifts from Dr Bert Vogelstein (Johns Hopkins University). HCT116 XRCC4 isogenic cells were purchased from Horizon Discovery. AT fibroblast cells (with and without ATM-reconstitution) were gifts from Dr Mukesh Nyati (University of Michigan). Cells were cultured in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% FBS, 2 mM glutamine and antibiotics. Cells were lysed with RIPA or NETN buffer containing protease and phosphatase inhibitors (Roche). Immunoprecipitation and western blotting were performed following standard protocol as described previously (21).