Fig. 1. Conserved non-coding sequences in the dac locus uncover two eye enhancers. (A) The dachshund genomic locus, with 5' and 3' eye enhancers indicated (see text for details). (B) AVID/mVISTA representation of the 5' eye enhancer (5EE). The numbered peaks indicate four areas of significant conservation. A small inversion in D. willistoni masks the first two peaks. (C) AVID/mVISTA representation of the 3' eye enhancer (3EE). The numbered peaks indicate six areas of significant conservation and two sub-fragments tested are shown. 3EE^{850 bp} contains all six CNCS blocks and is expressed only posterior to the MF. 3EE^{659 bp} contains the first four CNCS blocks and is expressed both anterior and posterior to the MF. The smallest active enhancer fragment identified is 194 bp (3EE^{194 bp}) and contains CNCS blocks 3 and 4. Regions of significant conservation are indicated in pink (B,C), and the predicted Sine oculis-binding site is highlighted in bright blue and is within CNCS block 3. (D,E) Representative third instar eye discs from 3EE^{850 bp}-GFP (D) and 3EE^{659 bp}-GFP (E) larvae triple labeled with GFP (green, D,E), Sens (magenta, D',E') and Dac (blue, D'',E''). GFP expression in 3EE^{659 bp} eye discs is detected anterior to the earliest Sens expression and overlaps with anterior Dac expression (E',E''). By contrast, GFP expression in 3EE^{850 bp} eye discs is not detected anterior to the anterior-most Sens-expressing column (D').