Craniofacial defects were quantified using linear measurements of specific cartilage elements as previously described (Figure 1d) (Everson et al., 2020 (link)). Whole-mount embryos were submerged in 50% glycerol 0.2% KOH in a 30 mm petri dish and imaged with an Olympus SZX7 microscope affixed with a DP22 digital camera. Whole body images were captured of embryos on their right sides at 3.2X magnification. Next, craniofacial elements were imaged both ventrally and dorsally at 5.6X magnification. Linear measurements of neurocranial elements were collected using ImageJ. The following measurements were gathered: whole body length (BL), whole skull length, trabecula length (TL), inter-trabeculae width (ITW) (i.e., distance between the trabeculae), and ethmoid plate length (EPL). Average measurements were compared between treatment groups using two-tailed ANOVA with post hoc Tukey's correction for multiple comparisons, with an alpha value of p ≤ .05 considered significant in Graphpad Prism v. 9.1.2.