Fig. 2. VP=Dach1 was misexpressed in the ectoderm by pouring the plasmid solution over the ectoderm. Diagrams at the top show the experimental design of electroporation. (A) A dorsal view of an electroporated limb bud on the right-hand side (Exp.) compared with a normal limb bud on the left. Expression of Fgf8 is obscure. (B) In a lateral view, Fgf8 expression is faint and blurred without making a clear boundary. (C) At stage 24, a severe depression was evident in the dorsal end of the limb bud. (D) Strong EGFP signals, which were derived from co-electroporated pCAGGS-EGFP, were observed in the ectoderm. (E) In such limb buds, Fgf8 expression was lost in the central part and deformed in the anterior (red arrowheads), whereas Fgf8 was normally expressed in the posterior (blue arrowhead). (F) Expression of Bmp7 was also lost in the central part (red arrowhead) and in the anterior mesenchyme, but was normal in the posterior area (blue arrowhead). (G) Mesenchymal expression of Fgf10 was repressed with inhibition of the limb outgrowth (red arrowheads). (H) Shh was expressed normally posteriorly (blue arrowhead). A depression similar to C was evident in the distal end (red arrowhead). (I) At stage 37, distal truncation was obtained. In the wing, where misexpression of VP=Dach1 was weak, truncation of digit II was evident. In the leg, where misexpression was extensive, distal autopod structures were completely missing (red arrowhead). (J) Alcian Blue staining of these limbs revealed a short digit II in the wing (red arrow) and the complete loss of digits in the leg (red arrowhead). Approximately 60% of the misexpressed limbs showed these phenotypes (n=242).