Single-Cell RNA Sequencing Analysis of Human Embryonic Stem Cell-Derived Neural Organoids

Human pluripotent stem cell-derived 3D neural organoids have been shown to recapitulate the major features and cytoarchitecture of early human brain development. We used three commercially available kits: STEMdiff™ Cerebral Organoid Kit, STEMdiff™ Dorsal Forebrain Organoid Differentiation Kit, and STEMdiff™ Ventral Forebrain Organoid Differentiation Kit. These neural organoids contain unique and diverse cell types which model the early brain and are patterned with developmental cues. Here, we sought to investigate the cellular diversity of these organoids using single-cell RNA sequencing. We generated dorsal forebrain, ventral forebrain, and cerebral organoids from H9 human embryonic stem cells (hESCs) that were subjected to single-cell dissociation on day 50 using an optimized dissociation protocol. We further utilized cell hashing to label multiple organoids (4 per condition) to explore intra-organoid variability within a culture.