Mouse anti brca1 d 9

The primary antibodies used were as follows: mouse anti-FLAG antibody (Sigma-Aldrich; F1804), mouse anti-γH2AX (Cell Signaling; #9718) and (Millipore; #05-636), rabbit anti-53BP1 (Novus Biologicals; NB100-304), mouse anti-53BP1 (BD transduction laboratories; 612522), rabbit anti-MDC1 (Abcam; ab11169), goat anti-Rif1 (N-20) (Santa Cruz, sc-55979), rabbit anti-beta-tubulin (Abcam; ab6046), mouse anti-c-Myc (Santa Cruz; sc-40), rabbit anti-H2AX (Cell Signaling; #2595). Secondary antibodies for western blotting were as follows: anti-rabbit igG, HRP-linked (Cell Signaling; #7074), anti-mouse IgG, HRP-linked (Cell Signaling; #7076). Secondary antibodies for IF analysis from Invitrogen were as follows: Alexa Fluor 488 (Rabbit, A11034), Alexa Fluor 594 (Rabbit, A11037), Alexa Fluor 594 (Mouse, A11032), Alexa Fluor 594 (Goat, A11058), Alexa Fluor 647 (Rabbit, A21245) and Alexa Fluor 647 (Mouse, A21236). For experiments involving cell cycle analysis, the following antibodies were used; mouse anti-BRCA1 (D-9) (Santa Cruz; sc-6954), mouse anti-RPA32/RPA2 [9H8] (Abcam; ab2175), rabbit anti-Phospho-histone H3 (Ser10) (D2C8) (Cell Signaling, #3377) and rabbit anti-CyclinA (H-432) and (Santa Cruz; sc-751).

Cell extracts were prepared and proteins were resolved by sodium dodecyl sulphate-polyacrylamide gel electrophoresis as described previously (47 (link),54 (link)). Rabbit anti-53BP1 were purchased from Novus and used at 1:1000 dilution. Mouse anti-BRCA1 (D-9, Santa Cruz) was used at 1:100 for western blotting. HA.11 clone 16B12 from Covance was used at 1:1000 for western blotting. Mre11 was used at 1:1000 for western blotting (Novus Biologicals). Secondary antibodies used were goat-anti-mouse IgG–horseradish peroxidase (HRP) conjugated, goat-anti-rabbit IgG–HRP conjugated at 1:2000 dilutions.