Act5C.CD2.Gal4 [49], and ck13 [29]. Marked clones of mutant cells were generated by Flp-mediated mitotic recombination subjecting flies once to a 35uC heat-shock for 30 minutes (Act5C,CD2.Gal4) or three times to a 38uC heat shock for 30 minutes (FRT42D Sans245). RNA in situ hybridization RNA in situ hybridization was performed as described previously [33]. The EST LD20463 was used as a template for RNA probe preparation. Western blotting Western blotting was performed as described previously [33]. Primary antibodies used were rabbit anti-Sans (1:1000; this study), rabbit anti-Cad99C, 1:1000 [42] and mouse anti-a-Tubulin, 1:1000 (Sigma T9026). Secondary antibodies used were HRP- conjugated goat anti-rabbit, 1:5000 (Santa Cruz sc-2054) and HRP-conjugated goat anti-mouse, 1:5000 (Pierce Biotechnology 31430). Immunohistochemistry Immunostaining of adult retinae was performed as described previously [50]. Ovaries were dissected and prepared for immunostaining using standard protocols. 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