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Briefly, whole Drosophila were fixed in 4% PFA for 12 h at 4 C, then rinsed 3 PBS for 3 h at RT with gentle shaking, and then immersed in FlyClear Solution-1 (8% THEED (2,2,2,2- (Ethylenedinitrilo)-tetraethanol) (Sigma-Aldrich), 5% Triton X-100 (Sigma-Aldrich) and 25% Urea (Fisher Scientific, in PBS) for 14 days at 37 C, as described previously 26

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