Fig. 5. Meis1 transcription factor is essential for CR4

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Fig. 5. Meis1 transcription factor is essential for CR4 Fig. 5. Meis1 transcription factor is essential for CR4.2-GFP expression.Analysis of homologous CR4.2 sequences from 11 species by Motif-based sequence analysis tools MEME (Bailey et al., 2006) revealed two highly conserved motifs (A,C). Meis1 transcription factor is essential for CR4.2-GFP expression.Analysis of homologous CR4.2 sequences from 11 species by Motif-based sequence analysis tools MEME (Bailey et al., 2006) revealed two highly conserved motifs (A,C). Electrophoretic mobility shift assays (EMSA) were performed using nuclear extracts isolated from E6 chick retina. Probe-3 (lane 2 in panel B) and Probe-5 (lane 2 in panel D) resulted in a band shift (arrowhead). This band disappeared with unlabeled competitors (lane 3); and the band reappeared with mutant competitors (lane 4). Mutant probes were synthesized with a 4 bp deletion of the TFBSs Hand (CTGG) (A) and Meis1 (TGAC) (C). (E–J) Chick retinas were injected and electroporated with a mixture of control CAG-DsRed (E,H) and a CR4.2-GFP mutant constructs (F,I) on embryonic day 4 (E4). Mutant constructs were generated by site directed mutagenesis. Transfected retinas were harvested and examined for GFP expression at E6. No change in reporter GFP expression was observed with deletion of Hand site (F); while no GFP expression was detected with deletion of Meis1 site (I). Scale bar: 1 mm. Mohammed M. Islam et al. Biology Open 2013;bio.20132279 © 2013. Published by The Company of Biologists Ltd