Fig. 1. Limb development in Daphnia. (A) First to fifth trunk limbs of Daphnia and their relationship to DLL expression. Epipod (Ep), exopod (Ex), endopod (En) and gnathobase (Gn) or filter comb (Fc) are arranged in a dorsal-to-ventral orientation. Endites of second to fourth leg bear prominent comb-like structures (Gn and Fc) that correspond to strong expression of DLL. No such branches or DLL expression are present in T1 or T5 (asterisks). (B,D,F) Daphnia embryos stained with nuclear marker YOYO-1. (C,E,G) DLL expression. Ventral view, anterior is upwards. (B,C) At 18 hours (Obreshkove and Fraser, 1940), with visible furrows separating thoracic segments, DLL expression begins in a dorsal cluster of cells in the first thoracic leg primordia, and in dorsal and ventral clusters in the second and the third primordia. Ventral DLL-expressing clusters in T2 and dorsal clusters in T3 are out of focus in this image. (D,E) At 21 hours, the DLL-expressing domain is expanded and clear nuclear localized signals are detected in future internal branches of developing thoracic legs and in first antenna, labrum, first maxilla, but not in mandibles and second maxilla. (F,G) Later stage embryos (24-27 hours) show strong DLL signals in limb branches, which show characteristic segment-specific morphologies. An1, first antennae; An2, second antennae; Lb, labrum; Mn, mandible; Mx1, first maxillae; Mx2, second maxillae; T1-T5, first to fifth thoracic segment; L1-L5, first to fifth thoracic limb. Scale bars: 100 µm.

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