Fig. 6. CR does not induce the dorsal position but induces the fan-shape arrangement of the ribs. (A-M) An ectopic CR (cr), obtained from a stage 14^- embryo, was labeled with DiI and grafted onto the host embryo dorsal to the endogenous CR (cr) at stage 13⁺. Observations were made of embryos 6 hours (stage 13⁺; A-D), 3 days (stage 15⁺; E-H) and 5 days (stage 17; I-M) after grafting. After 3 days of incubation, the grafted tissue integrated normally with the host tissue, and the expression of the CR-specific genes CRABP1 (G) and APCDD1 (H) was maintained in the graft. (J-M) Transverse histological sections at two different levels of the flank of the same embryo, as shown in I, stained with HE and Alcian Blue to visualize the cartilage. L and M are higher magnifications of the boxes in K. The ectopic CR is integrated completely into the host tissue and is associated with a thickened epidermis and underlying dense mesenchyme (L); the extracellular matrix is stained positively with Alcian Blue as the endogenous CR (M). Note in J and K that the pattern of rib growth has not changed on the experimental side and does not extend into the ectopic CR. (N) BrdU incorporation by the CR mesenchyme. BrdU was applied to a TK stage 16 embryo for 2 hours. Note the accumulation of BrdU at higher levels in the CR mesenchyme. (O) A schematic diagram showing the function of the CR in turtle embryos suggested by this study. In most amniote embryos (left, green), the ribs grow approximately parallel to each other compared with those in turtles (middle, red), which grow in a fan-like pattern resulting from the peripheral concentric growth of the carapacial primordium at the CR. Removing the CR arrests the growth of the carapacial primordium locally (right), and the anteroposterior growth of the vertebral column (purple arrow) causes the proximal parts of the ribs at the CR-ablated level (green) to become separated relative to the distal parts, resulting in the local disturbance of the distal parts of the ribs, as seen in Fig. 5M,N. Scale bars: 1 mm for A,E,I; 100 µm for B-D,F-H,N; 500 µm for J,K; 50 µm for L,M. fl, fore limb; hl, hind limb; m, myotome; n, notochord; nt, neural tube.