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Title: A molecular investigation of antero-posterior differences within the somite
Author: Hughes, D. S. T.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2007
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One factor implicated in mediating contact-repulsive activity is the molecule F-Spondin. However, findings with regard to its effects on the sensory neurite pathfinding are contradictory. Initially, I have shown that this contradiction results from membrane sequestration by COS7 cells of full-length protein used. The bulk of the work described here concerns the identification of genes differentially expressed between anterior and posterior half-sclerotomes as this is a prerequisite for the further investigation into the molecular mechanisms of somite segmentation and development. I developed a novel amplification method to generate sufficient material from anterior and posterior half-sclerotomes from mouse embryos to allow transcriptional profiling using oligonucleotide microarrays. Computational validation indicated that differences in expression data reflected differences between anterior and posterior sclerotome-halves. Statistical approaches identified a pool of ~60 candidates as putatively differentially expressed between anterior and posterior halves. Whole mount in situ hybridisation was used to verify the potential differential localisation of these candidates, resulting in 12-32 differentially expressed genes. Seven confirmed differentially expressed candidates were chosen on the basis of temporo-spatial expression patterns for further analysis using in vitro axon guidance assays. These secreted and transmembrane proteins were cloned into a mammalian expression vector such as to allow for production of soluble protein. The supernatants from transiently transfected cultured mammalian cells were used to conduct preliminary experiments into the potential of these molecules as axon repellents in the PNS.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available