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. Author manuscript; available in PMC: 2017 Nov 20.
Published in final edited form as: J Mol Biol. 2016 Oct 15;428(23):4608–4625. doi: 10.1016/j.jmb.2016.10.014

Figure 5. Expression of mcm10-m2,3,4 in budding yeast results in a severe growth defect and reduced RPA binding at replication origins.

Figure 5

A. 10-fold serial dilution analysis of budding yeast mcm10-1-aid cells expressing MCM10-WT, vector and mcm10-m2,3,4 from the GAL-S plasmid inducible promoter system (pRS415). Plates were incubated for 3 days at 25°C. B. mcm10-1-aid cells expressing MCM10-WT and mcm10-m2,3,4 were grown as described in Material and Methods. Cells were analyzed by FACS with propidium iodide staining for DNA content. C. mcm10-1-aid cells expressing MCM10-WT and mcm10-m2,3,4 were grown and chromatin immunoprecipitation was performed as described in Material and Methods. We used an antibody against RPA. Radioactive PCR bands were quantified, averaged and plotted. D. mcm10-1-aid cells expressing MCM10-WT and mcm10-m2,3,4 were grown as described in Material and Methods. D, upper panel. Mcm2 IP samples were analyzed by Western blot for expression of Mcm2. D, lower panel. Mcm2 IP samples were analyzed by Western blot for expression of phospho-Mcm2. Results from similar experiments were quantified, averaged and plotted. Graphs from (C) and (D) represent mean values from two independent experiments and error bars indicate the standard deviation of the mean.