Additional file 2.
Figure S5: RNA interference authenticates the Mcm10 protein in immunoblotting assays. (A) HeLa cells were transfected with GL2 or MCM10 siRNA and subsequently processed for immunoblotting with anti-Mcm10 antibodies (Ab [N] and Ab [FL]) to display the whole gel. The black and shaded arrow head points to the Mcm10 and a cross-reactive band respectively while NS refers to a non-specific band that displays equal protein loading. (B) Mcm10 levels after depletion of APC3, Cul1, Cdh1 and Cdc20. HeLa cells were transfected on three consecutive days with APC3, CUL1, CDH1, CDC20 or control GL2 siRNA and after the third transfection, asynchronous cells (ASN) were harvested for analysis of Mcm10 protein. NS points to a non-specific band that displays equal protein loading. (C) Mcm10 levels after depletion of beta-TRCP and FBXW7. HeLa cells were transfected on three consecutive days with BETA-TRCP, FBXW7 or control GL2 siRNA. After the third transfection, the cells were arrested with nocodazole for 15 h and were harvested after release from nocodazole (Noc) for analysis of Mcm10 protein, along with asynchronous cells (ASN). (D) The decrease of BETA-TRCP and FBXW7 mRNA was confirmed by RT-PCR. The numbers indicate the relative intensity of specific mRNA.
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Kaur et al. BMC Cell Biology 2010 11:84 doi:10.1186/1471-2121-11-84