PARP Trapping Effect Analysis

Immunoblot analysis was performed as described previously (16 (link)). For studying the PARP trapping effect of the PARPi, isolation of the nuclear soluble and chromatin bound fraction was performed using the Subcellular Protein Fractionation Kit for Cultured Cells (Thermo Fisher Scientific 78840) according to the manufacturer's protocol. Cells were therefore treated with 0.005% Methyl methane sulfonate (MMS, Thermo Fisher Scientific, 156890050) and 2 μmol/L talazoparib. The following primary antibodies were used for immunoblot analysis: anti-p21 Waf1/Cip1 (DCS60; Cell Signaling Technology 2946, 1:1,000), anti-pH2AX (Ser139; Merck, JBW301, 1:500), anti-β-Actin (Cell Signaling Technology 4967, 1:2,000), anti-p53 (DOI; sc-126, 1:500), anti-ATM (D2E2; Cell Signaling Technology 2873, 1:1,000), anti-PARP (Cell Signaling Technology 9542, 1:1,000), and anti-H2A.X (Cell Signaling Technology 2595, 1:1,000). The following secondary antibodies were used: anti-mouse IgG, horseradish peroxidase (HRP)-linked (Cell Signaling Technology 7076) and anti-rabbit IgG, HRP-linked (Cell Signaling Technology 7074).