Figure 1

Reduced cyclin B1 protein and kinase activity of Cdk1/cyclin B1. A. Upper panel: Breast cancer cells MCF-7, BT-474, SK-BR-3 and MDA-MB-231 cells were treated with cyclin B1 siRNA1, siRNA3, GFP siRNA (GFP) or without treatment (C) for 48 h. Cells were harvested and cellular lysates were prepared for Western blot analyses with antibodies targeting cylin B1, Cdk1 and β-actin. The later served as loading control. Lower panel: Quantification of cyclin B1 levels from Western blots (upper panel), normalized to β-actin. B. Kinase assay of Cdk1/cyclin B1 in vitro. BT-474 cells were treated with cyclin B1 siRNA1 or siGFP or without treatment (C). 24 h later cells were lysed and Cdk1/cyclin B1 complex was immunoprecipitated by using antibodies against cyclin B1 from cellular extracts. Normal IgG served as negative control. Kinase assays were carried out with the precipitates in the presence of histone H1 as substrate.