BMK primary cells were isolated and cultured as previously described (Degenhardt et al. 2002). Briefly, the kidneys from 4 to 5-d-old baby mice were minced with 1.5 mL of collagenase (0.1 U/mL)/dispase (0.8 U/mL) solution and incubated for 40 min at 37°C. After incubation, single cells were isolated from the solution and cultured in DMEM supplemented with 5% FBS, 2 mM L-glutamine, penicillin/streptomycin, gentamycin, and ITS liquid medium supplement (Sigma I3146).

In all experiments with BMK cells, the expression of the Mdm2I438K allele was induced by infection with recombinant adenovirus expressing empty vector control or Cre Recombinase for 24 h. After infection, the medium was changed and cells were left for 5 d. They were then treated with compounds (e.g., 4 μM Nutlin dissolved in DMSO for 8 h) and analyzed as indicated.

Recombinant adenovirus was purchased from the University of Iowa gene therapy core facility.

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