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Fig. 3. daf-15 and let-363 metabolic defect. Fat accumulation was assayed by Sudan black staining in L3 larvae grown at 20°C; daf-2 dauer larvae were formed constitutively at 25°C. (A) Wild-type N2, (B) daf-2(e1370), (C) daf-15(m81), (D) let-363(h111), (E) daf-16(mgDf47), (F) daf-16(mgDf47); daf-15(m81). All mutants except for the daf-16 single mutant accumulate abnormally high levels of fat relative to wild type. The entire experiment was performed twice, with at least 30 animals observed each time. Fat accumulation in daf-15(m81) and let-363(h111) was also confirmed by Nile red staining (data not shown).