Fig. 5. Whole-mount and sectional in situ hybridization analysis of Wnt-16 expression in the secondary palate.

For sectional in situ hybridization, 12 μm thick coronal frozen sections of embryonic heads on E12.5–E14.5 were prepared and adjacent sections hybridized with an antisense riboprobe (A–D) (for each gestation day) or a control, sense riboprobe (E–H) (for each gestation day). Representative sections were taken from various positions along the anterior-posterior (A–P) axis, as indicated by the corresponding line in panel (I). On E12.5, Wnt-16 was detected in palate mesenchyme along the entire A–P axis, with the expression domain becoming progressively restricted on E13.5 and E14.5. Black arrows indicate specific regions of Wnt-16 expression. These results were confirmed by an independent analysis on intact tissue (I,J) that demonstrated the temporally diminishing domain of Wnt-16 mRNA expression from E12.5 to E14.5. Only a small expression domain remained in the presumptive soft palate on E14.5. For orientation, the palatal shelves are outlined in panel J of each gestation day. All panels, 200X magnification.