In vitro experiment showed that HNF3beta was the direct regulator of sonic hedgehog (shh) promoter. To investigate the activity of zebrafish shh promoter in vivo, we constructed the expression vector pShh-EGFP with ligating a 538 bp zebrafish shh promoter,which contained two HNF3beta binding sites, to EGFP. The pShh-EGFP DNA was microinjected into one-cell stage embryos of the zebrafish and the embryos were observed for GFP expression with fluorescent microscopy. GFP expression started during gastrulation in the axial hypoblast layer. During segmentation, GFP was detected in the notochord but not in the foor plate. Our experiment demonstrated that the 538 bp shh promoter containing two HNF3beta binding sites is able to confer the notochord-expressing activity.
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