Supplementary Figure 8: Sensitivity of K4me2 clusters to technical aspects of the analysis.

A) The distribution of single cell coverage in each of the 4 clusters found and described in Figure 6A. B) The fraction of cells that were assigned to their original cluster when re-clustering a randomly chosen subset of 50% of cells (averaged over 100 random subsets). See also Figure 5C. C) The correlations between signatures are not driven by the overlap between them. The correlation between chromatin signatures based on co-variation across cells in the population is plotted vs. their overlap-based correlations, when using the real single-cell data (top) or shuffled data (bottom). Red dots indicate correlations between members of the chosen set of 91 signatures. See also the quality control for clustering analysis. The plots show that correlations in the shuffled data are equal to (and determined by) the overlaps between the signatures. In contrast, the correlations between the signatures in the real data are higher than that due to actual co-variation of signal in distinct genomic regions across single cells. D) The clustering of single cells is largely the same whether one uses the representative signatures, the entire set, or a completely different set of signatures. Top (replica of Fig. 5B): Multidimensional scaling (MDS) plot comparing the chromatin landscapes of single ES cells and MEFs (colored circles). The distance between any two cells is proportional to the distance between their 91-dimensional signature vectors. The plot shows 1,000 single cells (randomly sampled from the 5,405 cells with H3K4me2 data), colored by their cluster association. Middle: the same single-cell profiles re-clustered using the complete set of 314 chromatin signatures (see Online Methods). Bottom: the same single-cell profiles re-clustered using a different set of chromatin signatures (datasets taken from Meshorer, see Supplementary Note 1). E) The distributions of single-cell scores for G1/S related signature, G2/M related signature and OCT4 signature are plotted for ES1, ES2 and ES3. See Supplementary Note 1 for more information on the cell cycle related signatures.