Fig. 2. Downstream response to the induction of early embryonic signalling by Hex. (A) Real-time RT-PCR of Cerberus expression in VMZ explants analysed at stage 10.5. Embryos were injected with the indicated RNA and Cerberus expression quantified as in Fig. 1C. Data are based on three independent experiments. (B) In situ hybridisation for Cerberus expression and double in situ for Cerberus (purple) and nucGFP (light blue) expression at stage 10.5. Embryos were injected with 500 pg Hex, ß-catenin or both at the two-cell stage into both blastomeres (indicated with a schematic diagram in the lower right-hand corner) or into a single-ventral blastomere at the four-cell stage (indicated by `V') alongside a nucGFP RNA. Arrowheads indicate the site of injection. (C,D) Real-time RT-PCR analysis of Goosecoid and Chordin. Embryos were injected as in A and RNA from either animal caps (C) or VMZ (D) explants extracted and analysed at stage 10.5. Values are normalised to the expression level of Odc and the relative change in gene expression for the genes analysed was calculated by dividing the values from injected samples by the values from the uninjected. Data is based on three independent experiments. (E) In situ hybridisation for Goosecoid and Chordin expression at stage 10.5. Embryos were injected with 500 pg Hex, ß-catenin, or both, at the four-cell stage into a single-ventral blastomere alongside nucGFP. Double staining was performed. Arrowheads indicate the injected cells. High-magnification views of embryos co-injected with Hex and ß-catenin are shown for both Goosecoid (indicated as Gsc) and Chordin (indicated as Chd) expression. (F) Depletion of Hex by Hex MO. Embryos were injected as in Fig. 1F and in situ hybridisation for Cerberus, Goosecoid and Chordin performed at stage 10.5. Arrowheads indicate the ectopic domain induced by ß-catenin injection.