Basal characteristics of H1299 p53β and H1299 p53γ.

<p>(<b>A</b>) p53 immunoblot (Bp53-12) of H1299 cells transduced with p53β-tdTomato and p53γ-tdTomato construct. β-actin was used as loading control. (<b>B</b>) ³H-thymidine incorporation assay of H1299 p53β, H1299 p53γ and H1299 control cells (tdTomato vector control). The graph shows results from four individual experiments (average of 18 wells each). The experiments have been normalized. ** <i>P</i>-value<0.01. Error bars: Standard Error of the Mean (SEM). (<b>C</b>) Cell proliferation measured by Electric-Substrate Impedance Sensing. Impedance values are normalized after initial cell stabilization and shown as ratio of normalization value. The graph shows results from four measurements of vector control cells and six measurements of p53β⁺ and p53γ⁺ cells from two separate experiments. Standard error of mean is denoted by dotted lines. Highest variation in cell proliferation occurred after 30 hours after initiation, *<i>P</i>-value<0.05 calculated by paired Students t-test. (<b>D</b>) and (<b>E</b>) show immunoblot of basal level of Bax and p21, in both p53β⁺ and p53γ⁺ H1299 cells. GAPDH act as loading control, and the ratio of p21 or Bax to loading control with control value set to 1.0 is indicated. (<b>F</b>) Transient transfection of H1299 p53β and H1299 p53γ with the 13×p53RE-GFP construct and H1299 wt cells with both p53FL-construct and 13×p53RE-GFP were analyzed by flow cytometry (n = 2). <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056276#s2" target="_blank">Results</a> are presented as a ratio of GFP positive cells in H1299 p53β, H1299 p53γ and H1299 cells transiently transfected with full-length p53 to H1299 vector control. Error bars: standard error of mean. Student's t-test give <i>P</i>-value 0.053 of p53β cells versus vector control, and <i>P</i>-value 0.21 of p53γ versus vector control.