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. Author manuscript; available in PMC: 2006 Oct 1.
Published in final edited form as: Pigment Cell Res. 2005 Oct;18(5):349–359. doi: 10.1111/j.1600-0749.2005.00249.x

Figure 5.

Figure 5.

Cell cycle FACS analysis on HEK293 cells transfected with wild type (-) and (+) MITF WT. Twenty hours after transfection, cells were incubated with BrdU for 30 min and processed for BrdU and propidium iodide labeling before FACS analysis. (A) FACS plots of representative assays. Control represents non-transfected cells, and (-) MITF and (+) MITF correspond to transfections with the respective plasmids. The gates (R1-R10) were determined empirically. Note that HEK293 cells are aneuploid (hypotriploid) but undergo normal DNA doubling with each round of replication. R1, apoptotic, (hypo-)hypotriploid cells with no BrdU incorporated; and R2, apoptotic, (hypo)-hypotriploid cells that have incorporated BrdU. R6, apoptotic cells corresponding to (hypo-)hypohexaploidy. R3, G0G1; R5, G2M; R7, early S-phase; R8, late S-phase; R9, S-phase with low BrdU content; R10, S-phase with high BrdU content. Note that total S-phase corresponds to R9 + R10. (B-D) FACS analysis was repeated in three independent experiments, using the same gating as shown in (A). The results were quantified using the Cellquest software. (B) Quantitative analysis of the percentage of cells in G0G1, S, and G2M phases of the cell cycle. Note a significant difference between (-) and (+) MITF on S-phase (P < 0.02). (C) Separation of S-phase into low and high BrdU content. Note a significant difference between (-) and (+) MITF in low BrdU content (P < 0.01). (D) Separation of cells into early and late S-phase. Note a significant difference between (-) and (+) MITF on early S-phase (P < 0.01).