Fig. 2: Generation of kidney organoids through heterochronic recombination.

a Schematic illustration of protocol for heterochronic mixing, in which two directed differentiations are staggered by 2 days so that the first batch may be aggregated and cultured for 2 days before the second, newly differentiated, batch is added. b Representative stereo microscope image of kidney organoid generated through heterochronic recombination. c Immunofluorescence staining for Podocytes (Podxl), Proximal tubule (LTL), and distal tubule (Cdh1) in the organoid. d Integrated density for LTL, Cdh1, and Podxl in organoids generated from single batch (open bar) and heterochronic mixing (gray bar). Values calculated from n = 3 independent biological replicates and expressed as mean ± s.d. Unpaired t test with Welch’s correction was applied to calculate P value. e–i Representative high magnification immunofluorescence images of organoids derived from heterochronic recombination showing LTL⁺ Cdh1^- proximal tubule, Brn1⁺ Cdh1⁺ distal tubule, CDH1⁺ GATA3⁺ DBA⁺ connecting tubule or collecting duct, Podxl⁺ WT1⁺ Podocytes, CD31⁺ endothelial network, Pdgfrβ⁺ Meis1^- Pericytes (yellow arrow head), and Pdgfrβ⁺ Meis1⁺ stromal cells (white arrow head).