Fig. 5

Analysis of brain width and brain/placode boundary in sly mutants and control siblings. (A, A’) Immunostaining for HuC (cyan) at 36 hpf on Tg(cldnb:Gal4; UAS:RFP) (magenta) control and sly mutant embryos (frontal view). Similar immunostainings performed at 28 hpf are shown in Figure 5—figure supplement 1A, A’. (B, B’). Images of Tg(elementC:gfp); Tg(cldnb:Gal4; UAS:RFP) control and mutant embryos at 36 hpf (frontal view), similar images acquired at 28 hpf are shown in Figure 5—figure supplement 1B, B’. GFP (green) is expressed by the forebrain and a few OP cells. Arrows indicate where the brain width was measured (in three distinct positions along the DV axis). Measurements were also carried out at three distinct AP levels (through the z-stack). (C, D) Width of the forebrain in 36 hpf controls and sly mutants, at three different DV and three different AP levels (n=9 controls and n=8 mutants from four independent experiments, unpaired, two-tailed t test). Quantifications for the 28 hpf stage are shown in Figure 5—figure supplement 1C, D. (E-F’) Immunostaining for the OP marker Dlx3b (cyan) was performed on 36 hpf sly mutants and control siblings (frontal view). The signal was segmented using deep learning approaches (white signal), and the distortion index (see Materials and methods) of the OP/brain boundary was calculated in the regions outlined with red boxes. (G, H) Graphs showing the distortion indexes in controls and mutants at 36 hpf, for the left and right OPs (n=3 controls and n=3 mutants). ANOVA test (mixed models, with animals as random effect and genotype and side as fixed effects).