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Fig. 2 Positional cloning of the dae locus. (A) SSLP analysis places the dae mutation between two novel SSLP markers, scaf1747_9 (6/810 meioses) and scaf1747_7 (2/698 meioses) on linkage group 21. The fgf10 gene was subsequently identified as a candidate for dae. (B) The tbvbo allele has a K to stop mutation in amino acid 5, and the t24030 allele has an M to V mutation within amino acid 170 of Fgf10. (C,D) Injection of an Fgf10 morpholino directed against the exon2/intron2 splice acceptor site [e2i2 MO (red bar in B)] into wild-type embryos phenocopies the dae mutation. Injection of 0.125 mM morpholino (D) causes a severe truncation of the pectoral fin, identical to the phenotype seen in dae. (E) PCR amplification of the fgf10 open reading frame demonstrates splicing defects following morpholino injection (primer positions indicated in B). L, ladder; –, negative control; +, positive control; c, uninjected (compare with C); d, MO injected, i.e. dae-like phenotype (compare with D).