In an effort to test the feasibility of intracellular expression of enzyme-targeted antibodies to alter metabolism, recombinant antibody fragments in the single-chain format (scFv) were isolated from a phage display library using Arabidopsis chalcone isomerase (CHI) of the flavonoid biosynthetic pathway as the antigen. Each of the genes encoding the scFv's was cloned into a plant transformation vector, which was subsequently used to generate transgenic plants. One transgenic line with low expression of one of the scFv's appeared to have an altered flavonoid metabolism, as evidenced by a reduced capacity for anthocyanin accumulation and a reduction in flavonol glycosides in seedlings. Strong corroborating evidence that implicated the binding of scFv to CHI in the phenotypic alterations was obtained from protein mobility shift assays. Taken together, the results indicate that scFv-mediated metabolic alteration is possible in plants. Thus, we show that intracellular expression of scFv's can be exploited as an additional tool for metabolic engineering. / Ph. D.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/27349 |
| Date | 27 April 2001 |
| Creators | Santos, Michael Carmelo Orda |
| Contributors | Biology, Winkel, Brenda S. J., Beers, Eric P., Stevens, Ann M., Rutherford, Charles L., Walker, Richard A. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Dissertation |
| Format | application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
| Relation | msantos0401.pdf |
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