Identification of stage-specific KRAB-ZFP genes and validation by RT-qPCR.
A) Heat maps representing relative NanoString counts (blue = lowly/not-expressed, red = expressed) of genes with expression significantly associated with undifferentiated pluripotent cells (up), with differentiated/adult cell types (bottom), or that are similarly expressed in all different cell types (housekeeping-like genes, center), based on a non-parametric t-test. KRAB-ZFP genes are denominated using Ensembl gene IDs. Control genes that belong to either one of the three categories are included using their gene symbols. Arrowheads indicate the Ensembl gene IDs of the subset of genes, the nCounter expression level of which was validated by RT-qPCR. Green stars indicate KRAB-ZFP genes that are conserved in human. B) NanoString nCounter vs RT-qPCR expression analysis of a subset of KRAB-ZFPs. Values indicate average fold change of the selected KRAB-ZFPs in pluripotent cells (ESCs 2i+LIF/FCS+LIF or EGCs 2i+LIF/FCS+LIF) and differentiated cells (EpiSCs and EBs D6), expressed in function of ESCs FCS+LIF (set as 1). Fold changes were calculated as the average of three independent experiments. Error bars represent standard deviation values (SD) over the three replicates.