A new family of Drosophila balancer chromosomes with a w- dfd-GMR yellow fluorescent protein marker

Abstract

We report new w- fluorescent balancers scorable from stage 13 through adulthood that bear a nuclear-localized yellow fluorescent protein marker directly driven by dfd and GMR enhancer elements. The utility of this marker is enhanced by identification of an anti-GFP/yellow fluorescent protein (YFP) serum that is compatible with heat fixation.

Figure 1.—

dfd-based YFP constructs are effective markers for mid- to late-stage embryos. When viewing embryos by standard fluorescence microscopy, dfd-based YFP markers (A1–A4, B1–B4) are more visible than the commonly used actin-based GFP marker (J.-M. Reichhart and D. Ferrandon, personal communication) (C1–C4) using either GFP or YFP filter sets (Chroma Technology, Rockingham, VT). dfd-driven YFP fluorescence is readily visible in the head regions (asterisks in A1, A3, B1, and B3) while the more diffuse actin-driven GFP is marginally visible at stage 13 (arrows in C1 and C3) and moderately visible by stage 16 (arrow in C3). Note the nuclear localization of the nvYFP marker compared to the cytoplasmic localization of the eYFP and GFP markers (insets B3 vs. A3 and C3). In immunohistochemically stained embryos, the dfd-based YFP markers are also much more readily detected than the actin-GFP marker (A5 and B5 vs. C5). The shown embryos were heat-fixed (Miller et al. 1989; Peifer et al. 1993), stained with unpreadsorbed ab290 anti-GFP primary antibody at 1:10,000, and briefly developed to simulate use of the markers with antibodies that give moderate background so as to demonstrate the greater scorability of the YFP-based markers. Primary staining was visualized using biotinylated anti-rabbit secondary (1:300; Jackson Immunoresearch, West Grove, PA) and streptavidin–HRP (Vector Labs, Burlingame, CA) and developed using H₂O₂/diamino benzidine (DAB) (Sigma-Aldrich, St. Louis) for 45 sec. Images were acquired on a Zeiss Axioplan2 using an AxioCam at the same settings and subjected to the same linear level adjustments.