Fig. 2. Ectopic Ptc1 expression is localised to diastema mesenchyme. (A-D) E11.5 mandibular mesenchyme cultured for 24 hours. (A) Barx1 marks the molar regions. (B) Double-labelled (digoxigenin-fluorescein) in situ hybridisation marks Ptc1 (blue staining) diastema regions as distinct from Barx1-expressing (red staining) molar regions. (C) Adjacent sections through Ptc1-expressing domain (upper) demonstrate an absence of Barx1 (lower). (D) Adjacent sections through Barx1-expressing molar region (upper) demonstrate an absence of Ptc1 (lower). (E-H) Ectopic expression of Ptc1 and Gli1 in the diastema is due to Shh signalling. E11.5 mandibular mesenchyme cultured for 24 hours in the presence of 2H3 control antibody (E,G; normal Ptc1, Gli1 expression, respectively) or 5E1 Shh-blocking antibody (F,H; loss of Ptc1, Gli1 expression, respectively). (I,J) E11.5 mandibular explants cultured from Pro1(II)-lacZ transgenic mice demonstrating lacZ expression in Meckel's cartilage. (I) In the presence of epithelium Meckel's cartilage extends as two symmetrical rods. (J) In the absence of epithelium Meckel's cartilage extends further distally (arrowed), but no ectopic cartilages are visible after 24 hours. (K,L) Ihh expression is absent at E11.5 in the mandibular arch in both the presence (K) and absence (L) of epithelium after 24 hours. Scale bars: in A, 600 µm for A,B,E-L; in C, 600 µm for C,D.