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Title: Identification of the Prox gene family and the development of optical projection tomography for use in zebrafish
Author: Bryson-Richardson, Robert James
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2003
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Vertebrate homologues of the Drosophila melanogaster gene prospero have been identified in many species. Whilst the function and regulation of prospero has been well studied in Drosophila the function and regulation of the homologous vertebrate gene, praxl, is not known. We describe the identification of the prox genes as members of a multigene family in vertebrates through the isolation of new members of the Prox gene family in zebrafish, Fugu rubripes, Tetraodon nigroviridis, mouse, and human. We examined the phylogeny of this new multigene family and we characterised the expression of these novel genes in zebrafish. Analysis of the expression of these genes identified the slow muscle as site of expression for prox 1 that did not overlap with the novel zebrafish Prox genes. Therefore, we studied the function ofprox 1 in the slow muscle using a combination of DNA, and morpholino injections. We demonstrate that prox 1 in not required for the specification of slow muscle as determined by the expression of markers of terminal differentiation. We also show that the medial lateral migration of the slow muscle is unaffected by the loss of prox 1. However, ectopic expression of prox 1 specifically in the fast muscle causes a defect in nuclear patterning. In normal development the fast muscle cells fuse early to form a multinucleate syncytium. The nuclei in this syncytium are normally evenly spaced. Ectopic expression of prox 1 resulted in the nuclei of the fast cells being positioned at the centre of the syncytium similarly to the situation observed in the mononucleate slow muscle. Furthermore loss of Prox 1 results in the disrupted patterning of the slow fibres, demonstrating a role for Proxl in the patterning of the slow muscle fibres. An understanding of the 3-dimensional (3D) pattern of gene expression can often lead to a better understanding of gene function. Optical projection tomography (OPT) is a new method for obtaining 3D data about an object. OPT generates a 3D digital model of a sample and allows it to be virtually sectioned, or rendered to produce a 3D image. OPT was developed for use on mouse embryos and had not been tested with zebrafish. We describe the difficulties of using OPT on samples as small as zebrafish embryos and the development of techniques to overcome these problems and allow its use in zebrafish.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available