Figure 5

RNAi knockdown of DAE expression. The specificity of the DAE antibody was tested by knocking down its expression with RNAi. The midgut-specific Mex-Gal4 driver was used to induce RNAi. In the cross scheme used RNAi-expressing larvae were distinguished from non-expressing siblings by the presence of a GFP reporter in the latter. Larvae were sorted by GFP expression and then dissected and stained with the anti-DAE antibody followed by Texas Red labeled secondary antibody. Control larvae carrying the GFP-marked balancer chromosome exhibited the expected pattern of interstitial cell DAE staining (A) with the GFP reporter fortuitously expressed in neighboring copper cells (B). Siblings that expressed UAS-RNAi (recognized by lack of GFP; E) showed no detectable DAE staining (D), indicating that the antibody was specific for DAE. The merged image (F) was overexposed to demonstrate the presence of the middle midgut. Bar = 20 um.