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. 2006 Nov 10;34(21):6256–6263. doi: 10.1093/nar/gkl755

Figure 5.

Figure 5

M1 retention in Drosophila somatic tissues expressing UAS-MycTra2. (A) A schematic diagram is shown of the strategy for producing Tra2 overexpressing clones. Induced expression of yeast FLP recombinase from a FLP transgene driven by the hsp70 promoter causes fusion of ubitquitously active actin 5C promoter with Gal4 coding sequences in random cells. Clones of GAL4 positive cells are detected by activation of UAS-GFP and also express UAS-MycTra2. These clones were generated in flies also carrying a Tra2-β-galactosidase reporter transgene (CZP-ORF3) which produces RNA ubiquitously, but only expresses β-galactosidase if the M1 intron is retained. The reporter contains a frameshift mutation blocking translation from open reading frames used when the M1 intron is removed and initiating in exons 2 or 3. Expression of the reporter protein in intron-retaining RNA results from translation initiation at an AUG codon located downstream of M1 in exon 4. Shown are results from whole mount immunofluorescent staining of a larval imaginal disc (B, D and F) or brain tissue (C, E and G) that include clones of cells overexpressing UAS-MycTra2. Green staining (GFP) marks cells in which UAS-Myc-Tra2 is activated by Gal4 (B and C) and Red staining (β-galactosidase) marks cells with increased M1 retention (D and E). A merge of the red and green channels is also shown (F and G). Arrows in (D and E) indicate examples of large areas where GAL4 and Myc-Tra2 are not expressed and the reporter is not induced. Note that GFP staining is cytoplasmic and the Tra2-β-galactosidase fusion protein from the reporter is nuclear.