(a) Normalized expression levels of different PRKAR genes and a selection of reporter genes in embryonic stem cells (ESCs), neural progenitor cells (NPCs), and neural crest cells (NCCs), based on RNA-Seq data from different sources.23,24,25 For each individual set of expression data (ESC, NPC, and NCC), 0% reflects the gene with the lowest, and 100% the gene with the highest level of expression. The median expression level of each data set is 50% (dashed line). Genes scoring higher than 50% can be considered to be more highly expressed than the majority of genes in their respective data set. (b) Upper row: sagittal section of a human embryo at Carnegie stage 22 and corresponding 3D model of the embryonic brain (yellow: mesencephalon; green: subpallium; light blue: diencephalon; purple: hypothalamus; pink: rhombencephalon). A RNAscope PRKAR1β probe has been used to hybridize PRKAR1B messenger RNA (mRNA) (red). The section has been counterstained with hematoxylin (blue). A corresponding positive and negative control is shown in Fig. S2. Lower row: magnified sections show PRKAR1B expression in the pituitary, diencephalon, mesencephalon, and hypothalamus.