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Title: Stem cell pluripotency
Author: Hunter, Susan MacLean
Awarding Body: Cardiff University
Current Institution: Cardiff University
Date of Award: 2008
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Embryonic stem cells (ES cells) are derived by explantation of the embryonic portion of the pre-implantation embryo into culture. These cells have unique properties which have made them invaluable in study of the function of genes in vivo and of cell differentiation in vitro. They can be grown in culture for extended periods of time in an undifferentiated state and induced to differentiate in vitro. While undifferentiated they can be genetically manipulated. Subsequent reintroduction of these cells into the blastocyst results in the cells being integrated and contributing to all the cells of the animal including the germ line thus leading to designed genetic change. The homology of these cells, however, to their tissue of origin is not unambiguous. The primary aim of this thesis was to apply global transcriptome analysis to investigate the homology of ES cells to the pluripotent compartment of the embryo. Although ES cells can be grown in bulk, the tissue of origin, the embryonic portion of the peri-implantation embryo are small and inaccessible. It was therefore necessary to develop methods which would allow the transcriptome to be amplified without distorting the transcript profile. A linear amplification method proved to give the best result. The best method for fluorescently labelling the cDNA was shown to be enzymatic incorporation of aminoallyl dUTP followed by coupling to monoreactive Cy dyes. With these tools it was then possible to amplify the transcriptome of both colonies of ES cells and the embryonic portion of various peri-implantation embryos and apply the labelled cDNA to microarray slides. Statistical analysis of the results proved that the transcriptome of ES cells most resembles that of the embryonic ectoderm on day 5.5 of development.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available