Clonogenic cell survival assays after mitomycin C (MMC; Sigma) were performed, as described (Maranon et al., 2020 (link)). To assess cellular sensitivity to olaparib (AZD2281; Selleck Chemicals), cells were chronically exposed to 0.5–4 μM olaparib in regular growth medium for 12–14 days, as described (Spies et al., 2016 (link)). Cells were fixed and stained with crystal violet to determine the fraction of cells surviving.
Western blot analyses were performed according to our standard protocols (Wiese et al., 2006 (link)). The following primary antibodies were used: α-RAD51AP1 (Dray et al., 2010 (link); 1:6,000), α-RAD54L (F-11; sc-374598; Santa Cruz Biotechnology; 1:1,000); α-RAD51 (Ab-1; EMD Millipore; 1:3,000), α-PARP1 (ab6079; Abcam; 1:2,000), α-β-Actin (ab8226; Abcam; 1:1,000), α-Tubulin (DM1A; Santa Cruz Biotechnology; 1:3,000), α-HA.11 (MMS-101R; BioLegend; 1:1,000), α-Histone H3 (ab1791; Abcam; 1:10,000) and α-RAD54B (Wesoly et al., 2006 (link); 1:1,000). HRP-conjugated goat anti-rabbit or goat anti-mouse IgG (Jackson ImmunoResearch; 1:10,000) were used as secondaries. Western blot signals were acquired using a Chemidoc XRS+ gel imaging system and ImageLab software version 5.2.1 (BioRad).
Western blot analyses were performed according to our standard protocols (Wiese et al., 2006 (link)). The following primary antibodies were used: α-RAD51AP1 (Dray et al., 2010 (link); 1:6,000), α-RAD54L (F-11; sc-374598; Santa Cruz Biotechnology; 1:1,000); α-RAD51 (Ab-1; EMD Millipore; 1:3,000), α-PARP1 (ab6079; Abcam; 1:2,000), α-β-Actin (ab8226; Abcam; 1:1,000), α-Tubulin (DM1A; Santa Cruz Biotechnology; 1:3,000), α-HA.11 (MMS-101R; BioLegend; 1:1,000), α-Histone H3 (ab1791; Abcam; 1:10,000) and α-RAD54B (Wesoly et al., 2006 (link); 1:1,000). HRP-conjugated goat anti-rabbit or goat anti-mouse IgG (Jackson ImmunoResearch; 1:10,000) were used as secondaries. Western blot signals were acquired using a Chemidoc XRS+ gel imaging system and ImageLab software version 5.2.1 (BioRad).
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