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. 2011 Aug 30;6(8):e24312. doi: 10.1371/journal.pone.0024312

Figure 4. Effect of MAD2 knockdown on the development of nocodazole (Noco)-induced prometaphase arrest in MCF-7 cells.

Figure 4

A. Cells were transfected with siRNA MAD2 (si-MAD2) and the negative control siRNAs (si-Con), and 24 h later, cells were exposed to 250 nM nocodazole for additional 12 h. Then the whole cell lysates were analyzed for Cdc2, cyclin B1, MAD2, and Cdc20 levels using Western immunoblotting. B. The relative protein levels of Cdc2, cyclin B1, MAD2, and Cdc20 are calculated according to their densitometry readings, which are normalized according to the corresponding readings for the GAPDH protein bands. Each value is mean ± S.D. from three replicate measurements. * P<0.05 versus vehicle-treated control; # P<0.05 versus nocodazole treatment. C. Cells were transfected with si-MAD2 or siRNA negative control and then further treated with nocodazole (250 nM) for 12 h. Cells with Cdc2 knockdown were analyzed using immunofluorescent staining. Representative photographs were taken using a fluorescence microscopy (original magnification, ×200). D. Quantitative data on prometaphase-arrested cells. Each bar is a mean ± S.D. value from three separate experiments. ** P<0.01 versus the vehicle-treated control; # P<0.05 versus nocodazole treatment. E. The DNA content of cells was analyzed using flow cytometry as described in the Material and Methods section.