Flavonoids are plant secondary metabolites that are integral to our lives. Grapefruits are well-known for production of unique glucosylated products and the enzymes responsible are UDP-glucose:glucosyltransferases (GTs). The objective of this research was to obtain full-length clones of putative grapefruit GTs, express them, and characterize them. Previously, gene specific primers (from conserved PSPG box) and clone specific primers (from partial 5' clones) were designed, and a compiled sequence attained using SMART RACE RT-PCR. A full-length clone was obtained using primers designed from the extreme ends of the compiled sequence. The full-length clone was inserted into expression vector (pET32a) and transformed into expression host BL21(DE3)RIL. Expressed protein was tested for GT activity using different flavonoid aglycones and UDP-14C-glucose as glucose donor. Results indicated that the expressed protein was probably not a flavonoid GT. A directionally cloned grapefruit leaf cDNA library is undergoing EST mining to identify additional GT candidates.
| Identifer | oai:union.ndltd.org:ETSU/oai:dc.etsu.edu:etd-2065 |
| Date | 01 August 2004 |
| Creators | Roy Sarkar, Tapasree |
| Publisher | Digital Commons @ East Tennessee State University |
| Source Sets | East Tennessee State University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Electronic Theses and Dissertations |
| Rights | Copyright by the authors. |
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