Additional file 5.
Laser scanning cytometry. Exponentially growing BJ1 were fixed and stained for Mcm6, PCNA, phospho-S10-histone H3, and DNA as described in Methods. The volume of the staining reaction is greater than flow cytometry samples, but is otherwise the same. These data verify that both signals are nuclear (green = PCNA and orange = Mcm6). Mitotic cells were gated (not shown) and color-coded magenta. The pixels representing the rare mitotic events were made larger in Adobe Photoshop so that they would stand out. These data also support the notion that a more sophisticated analysis of bound Mcm6 versus PCNA might segment G1 into three, rule-based states, G1a1 [G1a cells with the lowest expression of Mcm6 and PCNA, equivalent to mitotic cells] (B); G1a2 [G1a cells (green) expressing Mcm6 at significant but sub-maximal levels] (C), and G1p cells (cyan), defined as the Mcm6 level at which cells enter S (D). Late S phase cells (red) are shown for visual comparison (E).
Format: TIFF Size: 3MB Download file
Frisa and Jacobberger BMC Cell Biology 2010 11:26 doi:10.1186/1471-2121-11-26