Extended Data Fig. 5. ETiX embryoids show neural folds and a developing tail bud with comparable differentiation trajectory and timing.
a. Day 7 ETiX embryoid recovered after stationary culture stained to reveal SOX2, SOX1 and DNA highlighting formation of the rostral neural folds (n = 11 ETiX7 from 4 independent experiments, n = 3 E8.0 natural embryos). b. Dorsal views of day 7 ETiX embryoid recovered after stationary culture (left) and natural E8.0 embryo (right) showing formation of SOX1 positive neural folds and BRY-positive notochord and tail bud (n = 11 ETiX7 from 4 independent experiments, n = 3 embryos). Scale bar, 100 μm. c. Lateral view of day 8 ETiX embryoid (top) and E8.5 natural embryo (bottom) showing FOXG1 expression in the telencephalon and OTX2 restricted to the forebrain and midbrain (n = 4 ETiX8 from 3 independent experiments, n = 2 embryos). Scale bar for a to c,100 μm. d. Quantification of brain area in E8.5 natural embryos and day 8 ETiX-embryoids. OTX2 was used to delineate the measured area. Data presented as violin plots with median and quartiles, each dot represents a sample (n = 6 E8.5 embryos and n = 17 ETiX8 from 7 independent experiments). Data are presented as violin plots with median and quartiles. Two-sided unpaired t-test, ns = p > 0.05 (p = 0.5223). e. Velocity plots for epiblast, neuroectoderm, surface ectoderm for all time points analysed in the inDrops sequencing dataset. f. Latent time analysis for epiblast, neuroectoderm, surface ectoderm for all time points analysed in the inDrops sequencing dataset. g. Quantification of the latent time analysis for epiblast, neuroectoderm, surface ectoderm for all time points analysed in the inDrops sequencing dataset