Figure 4: Transcriptional landscape of forward programmed MKs.

The undifferentiated hiPSC#1 line (n=2), day 20 fopMKs (hiPSC#1–2, n=4,2), day 25 ddMKs (hiPSC#1, n=4), day 10 cbMKs (n=4; all three groups CD42b+ sorted >95%) and day 30–69 LT-fopMKs (hiPSC#1 and #3, n=8 and 6; >80% CD42b+), were analysed for gene expression using Illumina Human HT-12 v4 BeadArrays. (a) Top five enriched gene ontology biological processes for differentially expressed (DE) genes in all MK samples compared to hiPSCs. −Log₁₀ P values are shown as colour scale. (b) Gene set enrichment analyses for DE genes from the different MK samples (versus hiPSCs; grey circles) against a ‘MK versus other blood types’ gene expression data set (from Haematlas²³). NES, normalized enrichment score; FDR, false discovery rate. (c) Gene expression correlation scatter plots on the whole-gene set using pairwise comparisons of different MK groups. R² Pearson correlation value and differentially expressed gene numbers are indicated. The differential expression threshold (two-fold-change; FDR 5%) is shown as dotted red lines. (d) Venn diagrams recording DE genes (|Log2 fold-change| >1; FDR5%) in hiPSC-derived MKs compared with cbMKs. The number of DE genes is indicated for each intersection and the top five enriched gene ontology term biological processes from the Venn Diagram intersections are shown. (e) Hierarchical clustering using the average agglomerative method on whole-gene data set. (f) Three-dimensional plots of the principal component analysis of MK populations. The first 3 PC are shown with respective percentages of variance indicated in brackets. The two principal component analysis boxes are snapshots of a rotation along the PC3 axis with MK sample groups highlighted.