Fig. 5: WNT signaling instructs the mesodermal lineage specification.

a Somitoid protocol with altered CHIR concentrations during the initial 2 days. The original protocol uses 8–10 μM CHIR. b Bright field (BF), Phalloidin staining, and ZO-1 staining images of the day 7 somitoids created with different CHIR concentrations. BF samples are different from Phalloidin- and ZO-1-stained samples. Scale bars: 350 µm. All samples stained (N = 3 (5 µM), 7 (6-7 µM), and 5 (8–10 µM)) showed similar expression patterns. c Classification of the day 7 somitoids created with different CHIR concentrations. Sporadic somites mean only 1–2 isolated somites, whereas strings of somites mean more than 3 rows of somites. N = 113 (5 µM), 62 (6 µM), 126 (7 µM), 46 (8 µM), 45 (9 µM), and 150 (10 µM). 4–12 independent experiments. d Quantification of the number of somite rows in the day 7 somitoids created with different CHIR concentrations. Mean ± SEM. N = 112 (5 µM), 56 (6 µM), 77 (7 µM), 14 (8 µM), 14 (9 µM), and 49 (10 µM). 4–12 independent experiments. Only the images with the entire somitoid structures were measured. c, d Part of samples of 8–10 µM is common to Fig. 1c, e. e HCR images of day 6 somitoids. Scale bars: 150 µm. f Volume quantification of lineage marker expression domains using HCR images of day 6 somitoids. SOX2 + & BRACHYURY-: Neural; UNCX4.1 + : Somite; BRACHYURY + & SOX2-: PSM; BRACHYURY + & SOX2 + : NMP. All samples stained (N = 3 (5 µM), 4 (6-7 µM), and 6 (8-10 µM)) showed similar patterns as long as somites were present. Microscopes: Opera (BF of panel b) and Light-sheet (Phalloidin and ZO-1 of b, e). Source data are provided as a Source Data file.