Figure 3: Fidelity of cloned cardiac SP cells to freshly isolated SP cells

(a) Pearson correlation plot, each cell of the heatmap showing the sample pair’s correlation coefficient (r). The clustering algorithm separates highly correlated samples (red: r close to 1) from weakly correlated ones (blue: r close to 0). The 20 independent lines of cloned cardiac SP cells show strong within-group correlation, moderate correlation to MSCs and ESCs, and weak correlation to other samples including heart. All clones were analysed at <30 passages. BM, bone marrow; CM, neonatal cardiomyocytes; eH, embryonic day 10 heart; ESC, undifferentiated AB2.2 ESCs; H, adult heart (encompassing whole heart, atria, and each ventricle); K, kidney; L, liver; MSC, PDGRα⁺ bone marrow MSCs; Sk, skeletal muscle; Sp, spleen. (b) Molecular signature of cloned cardiac SP cells. Bar graph, mean±s.e.m. for all 40 genes’ expression in 20 independent clones. Genes are colour coded based on functional association or tissue specificity. For comparison with reference samples, see Supplementary Fig. 3. (c) Density plot showing the prevalence for expression of key cardiac transcription factors in the 20 clones. See Fig. 1d. The clustering algorithm separates genes with heterogeneous (multimodal) expression from those with homogeneous low or high expression. (d) Heatmap showing scaled expression of the four most heterogeneous transcription factors in the 20 clones (Gata4, Mef2c, Tbx5, Hand2 and Nkx2-5). Expression within each sample is scaled to Z-score, 0 indicating mean expression of the four genes. Red indicates higher expression than the mean, while blue indicates lower expression. The two-dimensional hierarchical clustering algorithm orders the clones and genes based on co-expression profiles. Asterisks denote four clones taken forward to more detailed studies. (e) Western blots for cardiac transcription factors in the clonal lines. Cytoplasmic (C) and nuclear (N) fractions were analysed using glyceraldehyde 3-phosphate dehydrogenase and histone H1 to authenticate the fractions and β-actin as a loading control. The bands for TBX20 correspond to isoforms with or without the C-terminal 145 a.a. extension.