Figure 1.

noxa and puma are not expressed in noxa^−/−puma^−/− double knockout mice. (a) RT-PCR analysis on cDNAs generated with total RNA from thymocytes and spleen cells from mice of the indicated genotypes. Wt thymocyte cDNA, which was used in a reaction without reverse transcriptase enzyme, is included as a negative control. The identity of the PCR products was confirmed by Southern blotting using an internal oligonucleotide specific for noxa cDNA as a probe (top panel). RT-PCR analysis with primers specific for hprt was used as a loading control. Sizes of DNA size standards (in bp) are shown on the left hand side (bottom panel). (b) Western blot analysis of thymocytes from wt, noxa^−/−, puma^−/− and noxa^−/−puma^−/− mice cultured for 7 h in the presence or absence of 1 μM dexamethasone or following 2.5 Gy γ-irradiation. Western blots were probed for Bim, Bid, Bad or Puma protein levels. Probing for β-actin was included as a loading control. The molecular weight (in kDa) of protein size standards is shown on the left hand side. (c) Western blot analysis of Puma protein expression in wt or p53^−/− thymocytes cultured without treatment (Unt) or following treatment with the indicated doses of γ-irradiation. Time in culture is indicated in hours. The molecular weight (in kDa) of protein size standards is shown on the left hand side