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. 2011 Sep 30;286(46):39776–39785. doi: 10.1074/jbc.M111.226464

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© 2011 by The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 4. — Overexpression of MCM3 blocks S phase entry. A, Tet-on inducible HeLa cells (MCM3 and MCM3 T722A) were treated with or without tetracycline for 24 h. Cell extracts were prepared and subjected to immunoblot with anti-FLAG antibody. B, Tet-on inducible HeLa cells for control, FLAG-MCM3, and FLAG-MCM3 T722A were treated with tetracycline. The chromatin-bound fraction and total cell lysates were prepared for immunoblotting with indicated antibodies. C, Tet-on inducible HeLa cells for control, FLAG-MCM3, and FLAG-MCM3 T722A were treated with tetracycline and subject to flow cytometry analysis. D, schematic view of experimental design. Tet-on inducible cells were treated with 2 mm thymidine for 16 h and released in fresh medium for 8 h and then treated again with 2 mm thymidine and tetracycline for 16 h. Cells then were released for 3 and 6 h and subjected to flow cytometry analysis. E, cell cycle profiles were analyzed by flow cytometry analysis.