Only emx3 and emx1 regulate expression of eomesa. A-E: Injection of emx3 morpholino antisense oligonucleotides (MO; D,E), to a lesser extent emx1MO (B), but not emx2MO (C), leads to reduced expression of eomesa compared with control embryos (A). F: The emx3 morpholino phenotype is not enhanced when emx1MO and emx2MO are coinjected. emx3 splice morpholinos (E) give rise to the same phenotype as translation blocking morpholinos (D), and lead to aberrantly spliced mRNA (H,I), which together confirms specificity of emx3 morpholino phenotypes. G-I: Reverse transcriptase-polymerase chain reaction (RT-PCR) of RNA from embryos injected with emx2 and emx3 splice morpholinos that target exon 2 leads to aberrantly spliced mRNAs. Exon1 is fused to exon3 of emx2 (G) and emx3 (H), respectively. Alternatively, the third exon of emx3 is spliced to a cryptic splice site within exon 2 that leads to an in-frame deletion of parts of helix 2 of the homeodomain (I). A-F: Frontal views of 24 hr embryos, dorsal to the top. Scale bar = 100 μm.
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