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FIGURE 1. Induction of hepatogenesis in Gata4-expressing AC explants and AC/AE conjugates. (A,B) Marker gene analysis in Gata4-expressing animal caps. (A) RT-PCR showing expression of cardiac fate marker myl7, liver fate marker nr1h5 and control marker odc1 in Gata4 mRNA-injected ACs, control ACs and stage 33 sibling control embryo. Representative of four independent experiments. (A′) Western blotting shows efficient translation of exogenous Gata4 protein. An equivalent of ½ embryo at st. 9 from a sample of five embryos has been loaded on the gel. (B) qPCR showing fold difference in expression between Gata4 treated animal caps and control animal caps grown to st. 33, induction of expression of myl7, nr1h5, hhex and marker of endoderm foxa2 is shown. Number of independent experiments (n) and standard error of the mean are shown. (C–E) Cardiac and liver domains are adjacent in AC/AE conjugates and (F) age matched sibling controls. AC/AE conjugates (sandwiches) were cultured until sibling control embryos reached st. 34 and were processed for myl7 expression (turquoise) and nr1h5 (purple) by WMISH. AC-derived tissue is shown in pink/red. Three AC/AE examples are shown demonstrating adjacent expression domains of myl7 in the AC-derived tissue and nr1h5 in the AE-derived component of the explant (12/12 specimens showed the same pattern). Dashed line highlights the border between AC and AE. |
