Gene Expression Analysis of hESC Differentiation Figure 5. Hierarchical clustering of 5955 differentially expressed genes was performed using the mean signal intensity for each replicate. Three biological replicates of hESCs and differentiated cells at the NI, NE, and DN stages were compared and showed show high intraclass correlations compared with interclass correlations. Five distinct clusters were distinguishable based on the expression patterns of the different cell types. The differentially expressed transcripts were clustered into five expression groups, including (A) 2576 genes that were upregulated in DNs compared to other stages; (B) 720 genes that were upregulated in NEs; (C) 326 genes that were upregulated in hESCs and NIs compared with NEs and PLoS ONE | www.plosone.org 7 July 2011 | Volume 6 | Issue 7 | e22856