Sei1 in Cancer and Diabetes Figure 4. Proliferation, immortalization and transformation of Sei1-null MEFs. (A) Growth curves of primary MEFs grown at the indicated serum concentrations. (B) Senescence and immortalization were evaluated performing a 3T3 serial passage protocol of MEFs of the indicated genotype. DPDL, increase in population doubling level (C) Colony formation assay measuring the immortalization potential of MEFs of the indicated genotypes. (D) Foci formation assay of MEFs transduced with HRasV12 or with bicistronic E1A/HRasV12. In all the above assays, at least three independent preparations of primary MEFs per genotype were used. Values correspond to average and s.d. doi:10.1371/journal.pone.0008744.g004 Figure 5. Apoptosis in Sei1-null cells. (A and B) Three independent preparations of primary MEFs per genotype were either irradiated with the indicated doses of UVC (A), or transduced with E1A and treated with low serum conditions or with adriamycin, as indicated (B). Apoptosis was measured by quantifying subG0 DNA content by FACS. (C) Thymocytes from three mice per genotype treated with 10 Gy of c-radiation were isolated and apoptosis was measured 3 h post-radiation, as in (A) and (B). (D) Protein extracts of lung and spleen from the same irradiated mice as in (C) were immunoblotted against p53 or its target p21Cip1. Actin was used as a loading control. doi:10.1371/journal.pone.0008744.g005 PLoS ONE | www.plosone.org 5 January 2010 | Volume 5 | Issue 1 | e8744