Supplementary Figure 3: Time series-based RNA sequencing captures developmental gene expression dynamics.

a, Representative images of retroviral lineage-tracing experiments in progressively maturing neurons. Retrovirus (RV)-labeled cells express eGFP and co-expression of PSA-NCAM is indicative of the neuronal lineage. Scale bar 20 μm. Immunohistochemistry was performed in all 13 subject lines and 2 cell culture replicates each with similar results (also see Supplementary Figure 5a). b, Schematic of FACS-based purification of NSCs and defined subpopulations of eGFP⁺/PSA-NCAM⁺ neurons over the time course of in vitro differentiation. c, Heatmaps of time-course RNAseq expression analysis showing the top 300 upregulated and top 200 downregulated genes (by fold change) during neuronal differentiation. Expression values are shown for one representative control cell line. Corresponding GO term analyses for differentially expressed genes (Wald test implemented in the DESeq2 package with Benjamini-Hochberg adjusted P < 0.01) are depicted in the table to the right (Supplementary Table 2 and 3); GO terms upregulated: n = 2220 genes, GO terms downregulated: n = 1344 genes.