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Title: A murine model for haemangioblast transplantation
Author: Babaie, Yasmin
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2004
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A promoter trap strategy was used to target a single allele of the flk-1 gene with one of two reporter constructs. One placed the EGFP reporter under flk-1 transcriptional control, while the other transcribed the selectable HPRT enzyme, which could be used to isolate flk-1 expressing cells in the HM1 hprt deficient ES cell line. Homologous recombination was successfully achieved with both targeting vectors. HPRT targeted selectable lines were successfully isolated. All lines showed appropriate transgene expression at high enough levels for effective HAT selection to isolate pure populations of flk-1 expressing cells. Two methods of deriving haemangioblastic progenitors from ES cells were tested for their efficacy of the isolation of a pure haemangioblast population based on the expression of flk-1. Methylcellulose cultures were found to be highly variable and problematic to carry out successfully whereas monolayer differentiation on collagen IV was more consistent. This system was then used to optimise the differentiation and selection regime applied to the flk-1/HPRT targeted cells in an attempt to isolate a pure haemangioblast progenitor population. Flk-1/HPRT targeted cells were marked with a constitutive GFP transgene, differentiated and selected for flk-1/HPRT expression on collagen IV. The selected cells were injected into the blastocyst stage embryo to see whether a developmentally more advanced cell population would survive and contribute to the expected cell lineages. Contribution of the GFP marked cells was observed in mesodermal and highly vascularised organs such as the heart, liver and kidney but was absent from the ectodermally derived brain. Expression was strongly localised around regions of vacularisation.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available