This project and the experiments discussed in this document sought to contribute a functional basis for SNPs within G6PC2 significantly associated with variations in fasting blood glucose by genome-wide association study. Through a combination of binding, reporter gene expression, and in vitro minigene splicing analyses, the roles of a number of SNPs were examined, including rs573225, rs13431652, rs2232316, and rs560887. It was demonstrated that rs13431652 and rs2232316 affect binding of NF-Y and Foxa2, respectively, with changes seen in fusion gene assays correlating these data with the genetic associations of the GWAS. Variations in Foxa2 binding were also observed with allelic variation at rs573225, although fusion gene analysis of this SNP failed to support the binding and genetic data, as well as the function of G6PC2. Because of this inconsistency, future utilization of cell lines and transient transfection for this nature of experiment must be heavily scrutinized.
Additionally, splicing variation of G6PC2 exons, based on allelic variation at a number of SNPs, was assessed. By utilizing a minigene plasmid capable of forming a spliced RNA product in vitro, it was found that rs560887 significantly affects splicing of exon 4. The magnitude of this result was found to be based in part on the non-consensus 5 exon 4 splice junction present in humans.
These experiments as a whole serve to corroborate and lend a functional basis to the SNPs of G6PC2 associated with variations in fasting blood glucose.
| Identifer | oai:union.ndltd.org:VANDERBILT/oai:VANDERBILTETD:etd-01252012-093840 |
| Date | 25 January 2012 |
| Creators | Baerenwald, Devin August |
| Contributors | Maureen Gannon, Roland Stein |
| Publisher | VANDERBILT |
| Source Sets | Vanderbilt University Theses |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.vanderbilt.edu/available/etd-01252012-093840/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to Vanderbilt University or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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