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Figure 5 SMYD3 depletion enhances early cardiac differentiation in mESCs. (A) Gene expression of early cardiac markers Isl1 and Nkx2-5 was analyzed by qRT-PCR. Data were normalized to Gapdh. Means and SEM from three independent experiments are shown. Significance was calculated by 2-way Anova, followed by Bonferroni post hoc test. ** p < 0.01, *** p < 0.001 value for the significance is shown in the plot. (B) Western blot analysis was performed on mESCs protein extracts at different time of differentiation, with antibodies raised against Flk1. GAPDH serves as a loading control. Normalized band intensity in immunoblots is reported below signals. (C) Immunofluorescence analysis was performed with Flk1 antibodies at d5 of EB differentiation. DAPI stains nuclei. Scale bar: 200 μm.