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Additional Supporting Information may be found in the online version of this article.

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DVDY_22734_sm_SuppFigS1.tif294KSupporting Information Fig. S1. Schematic of positive and negative intersectional methods for labeling subgroups of cells. A: Positive intersectional method: a group of cells is labeled by partially overlapping expression domains of two different enhancers. Expression occurs only in cells in which both components (for example, of split Gal4) are present. B: Negative intersectional method: a group of cells is labeled by two different enhancers; expression occurs in the cells in which only the positive activator is present.
DVDY_22734_sm_SuppFigS2.tif17570KSupporting Information Fig. S2. Subgroups of neurons can be distinguished by inhibiting Gal4-driven expression with a fluorescently tagged Gal80. Single slice confocal images, lateral views, anterior to left, dorsal up, of 72 hours postfertilization (hpf) eyes in Tg(isl2b.3:Gal4)zc65; Tg(UAS:GFP) embryos. Immunostaining for green fluorescent protein (GFP), green; TagRFP, red; Topro3 nuclear stain, magenta. A–A″′: Embryos injected with hsp70l:TagRFP (no Gal80 expression). B–B″′: Embryos injected with hsp70l:Gal80-2A-TagRFP and heat-shocked at 48 hpf show expression of TagRFP (RFP) and inhibition of Gal4-dependent expression. C–C″′: Embryos injected with hsp70l:Gal80-TagRFP and heat-shocked at 48 hpf show RFP expression and concomitant inhibition of Gal4-dependent expression. (A″″–C″″) shows magnified view of the insets in (A″–C″), demonstrating disjoint GFP and RFP expression. Scale bar = 50 μm.

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