Extended Data Fig. 3: Inhibition of acto-myosin activity does not interfere with anisotropic stress mediated development of EK formation.
(a) Schematic depicting the dissection, agarose embedding, injection of oil microdroplets, and imaging of the incisor. The image of the embryo and the microscope have been obtained from Biorender.com. (b) Outline of E13.5 incisor with reference points 1–3 illustrating the marker locations on the border and point 4 illustrating the oil droplet for vector field and microdroplet orientation analysis. (c) Anisotropy analysis of the live E12 and E13.5 murine incisor. (E12: n = 6 and E13.5: n = 6). (d) Representative E14.5 incisor epithelium separated from the mesenchyme after 45 mins dispase treatment. (n = 3) (e) EdU and E-Cad immunostaining after a 1 h EdU pulse chase in E12 murine incisors cultured with DMSO or blebbistatin for 40 h. Quantification of total EdU positive cells in each field per condition for all incisors. (Control: n = 7; Blebbistatin: n = 7) (f) Whole mount in situ of Shh and E-Cad immunostaining in E12 incisors cultured for 40 h with DMSO or Y27632 (ROCK inhibitor). (DMSO: n = 5; Y27632: n = 5). Scale bar, 25 µm. Dashed line outlines the incisor. Representative 2D central plane shown in all images. Data are represented as mean ± s.e.m. Statistical analysis was done using the unpaired two-tailed Student’s T test (assuming unequal variance) with Welch’s correction for e. n represents number of embryos; one incisor measured per embryo.
