Different constructs used to generate stable transgenic lines and corresponding distribution of EGFP reporter in expected olSix3.2 expression domains. (a) Drawings to the left of the panel are schematic representations of the different constructs (cI and cVI to cXXII) used to generate stable transgenic lines, whereas the tables to the right summarize the presence (+) or absence (-) of enhanced green fluorescent protein (EGFP) reporter expression corresponding to the expected olSix3.2 expression domain at different stages of differentiation, in the retina or ectopically in the spinal cord. The red box represents the 5'-untranslated region and the first nine nucleotides of the olSix3.2 coding sequence, in frame with a nuclear EGFP reporter, whereas the dark blue box represents the minimal tyrosine kinase promoter. (b to e) The images show frontal vibratome sections through the optic cup of in situ hybridized (b) wild type and (c) cVII, (d) cXVIII and (e) cXIX transgenic lines. Note that module D alone is sufficient to drive EGFP expression in the hypothalamus and neural retina but not in the lens (empty arrow in panel e), whereas in its absence EGFP expression is completely lost (panel b). A similar absence of EGFP expression was observed in the cVIII to cXVII transgenic lines, all of which lack module D. Note also that the combination of modules D to H is necessary for expression in the lens placode (arrow in panel d), as indicated by in situ hybridization of the endogenous olSix3.2 distribution (arrow in panel b). Hy, hypothalamus; NR, neural retina.