Differentiation and Single-Cell RNA-seq Analyses of Human Pluripotent-Stem-cell-derived Renal Organoids.

Here, we present a protocol for the maintenance and differentiation of human pluripotent stem cells into renal organoids. We describe steps for using a series of readily made differentiation media, multiplexed sample single-cell RNA-seq analysis, quality control, and validation of organoids using immunofluorescence. This provides a rapid and reproducible model of human kidney development and renal disease modeling. Finally, we detail genome engineering using CRISPR-Cas9 homology-directed repair for the generation of renal disease models. For complete details on the use and execution of this protocol, please refer to Pietrobon et al.1