Generation and Characterization of dfoxoΔ94 Allele

The dfoxo^Δ94 allele was generated by conventional imprecise excision using P[GT1]foxo^BG01018 flies that carry an P[GT1] element transposon in the 5′upstream region of the dfoxo gene, approximately 130 nucleotides upstream of the dfoxo transcriptional start site (Dionne et al., 2006 (link)). The 5′ and 3′ breakpoints of the dfoxo^Δ94 deletion were mapped to the genomic sequence by PCR and sequencing. UASp-dFOXO transgenic flies for germline expression of dFOXO were generated using standard procedures. The P[GT1]foxo^BG01018, daughterless-GAL4 (da-GAL4), UAS-InRDN (K1409A), UAS-Dp110DN (D954A), eyeless-GAL4 (ey-GAL4), and mata-GAL4 stocks were obtained from the Bloomington Stock Centre. daughterless-GeneSwitch (daGS) was kindly provided by Veronique Monnier (Tricoire et al., 2009 (link)). InsP3-GAL4 was kindly provided by Michael Pankratz (Buch et al., 2008 (link)). All stocks were backcrossed for at least 6 generations into the control white^Dahomey (w^Dah) stock. w^Dah was derived by backcrossing white¹¹¹⁸ into the outbred wild-type Dahomey background. Flies were raised and maintained on standard sugar/yeast medium (Bass et al., 2007 (link)). Stocks were maintained, and experiments were conducted at 25 °C on a 12:12 hours light/dark cycle at constant humidity.