Research Paper Volume 10, Issue 6 pp 1454—1473

Figure 7. PARylation of cell extract proteins of NMR and mouse cells in presence of [³²P]NAD⁺. (A) The activated DNA was incubated with 0.5 mg/ml of cell extracts of NMR cells (lanes 1 – 5) or mouse cell (lanes 6 – 10). The control probes (lanes 11–14) contained 70 nM PARP1 instead of cell extracts and 40 µM [³²P]NAD⁺ (lanes 11 and 13) or 400 µM [³²P]NAD⁺ (lanes 12 and 14). Lanes 13 and 14 correspond to lanes 11 and 12 but with low exposure. The reaction mixtures were incubated at 37⁰C for 4 min. The mixtures were supplemented with loading buffer, heated at 97⁰C for 10 min and then analyzed as described in the section ‘PARylation of cell extract proteins’. (B) Quantification showing the yield of PARylation products (A). Post-irradiation time is the time the cells were cultivated after irradiation before the cells were harvested for the following extract preparation. “n” designates non-irradiated cells.