| Dissertation |
Thesis (Ph.D.) --NUI, 2012 at Department of Anatomy and Neuroscience, UCC. |
| Summary |
Before the potential use of stem cells in transplantation can be realised, it is necessary to address the formation of tumours following such a procedure. We aimed to produce a construct which would allow for the selective ablation of tumourigenic cells following stem cell transplantation. Initially, a “suicide” gene construct was produced which would allow for selective ablation of proliferating cells following stem cell transplantation. This “suicide” gene construct consisted of a Top2A cell cycle-regulated promoter, cloned upstream of a loxP-flanked eGFP reporter gene and a diphtheria toxin fragment A (DTA) sequence. A “testing” construct was also generated, consisting of an RFP sequence in place of the DTA, which would allow for the initial testing of the construct's stability. Cell cultures which had been cell cycle-regulated were established to test the functionality of the Top2A promoter. A decrease in GFP expression from the “testing” construct following serum starvation, confirmed the functionality of Top2A promoter. Following confirmation of the promoter activity, Cre recombination studies on the “testing” construct indicated the successful recombination of this construct. With increases in Cre concentration and in time, increases were observed in the number of cells expressing RFP, while decreases were observed in the number of cells expressing GFP. This suggests that the number of fully-recombined cells will increase with time, to eventually allow the complete recombination of the “testing” construct. However, full recombination of the “suicide” gene construct may take longer to achieve, as was indicated by the extended experimental time and lack of decrease in co-expression of GFP and DTA in our studies. Based on the results of this study, it is clear that the use of a cell cycle-regulated DTA toxin construct in the ablation of tumour-forming stem cells may be a viable method for the prevention of tumour formation following transplantation. |
| Subject |
Stem cells.
|
|
Gene therapy.
|
| Collection |
Theses Ph.D.
|
|
Theses Anatomy and Neuroscience Department
|
| Description |
191 p. ; 30 cm. |
|
