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Fig. 1
The RH2-LCR is an evolutionarily conserved enhancer in teleosts. a Sequence comparison of the RH2 locus between zebrafish and medaka (M) is shown using mVISTA program. The baseline corresponds to the zebrafish RH2 region. An enlarged illustration around the RH2-LCR is also shown. Black and gray bars under the chart are the exons of the RH2 genes and the other genes, respectively. The red bar indicates the RH2-LCR. The sequence homology is indicated to the right of the chart. b Sequence comparisons of the RH2 locus of medaka with zebrafish (Z) and Tetraodon (T) are shown as in (a). Homology regions colored with gray correspond to coding regions of genes and those colored with pink correspond to conserved non-coding sequences. Note that Tetraodon has higher homology at the RH2-LCR than zebrafish, reflecting their closer relation with medaka. c Construction of the medaka RH2-A/GFP-BAC clones (upper panel) and expression levels at 5 dpf of the GFP reporter in zebrafish injected with the BAC clones above (lower panel). The histogram shows the percentage of eyes graded into four levels (+++, ++, + and -) according to the number of GFP-expressing cells in the retina. The names to the left of the histogram indicate the constructs injected. The numbers to the right of the histogram show the total number of eyes examined. d, e Whole mount retinas of 7-dpf zebrafish embryos injected with the RH2-A/GFP-BAC (d) and with mixture of the medaka RH2-LCR fragment and the GFP reporter under the 3-kb upstream region of RH2-A. GFP fluorescent signals appear as green (left) and immunostaining signals of SDCs by the anti-RH2 antibody appear as magenta (middle). Overlap of the two signals appears as white. The right panels are the overlays of the left and middle panels. Scale bars = 10 µm