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Browsing by Author "Broeckling, Corey D., author"

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    Metabolic engineering of Arabidopsis for butanetriol production using bacterial genes
    (Colorado State University. Libraries, 2013-11) Abdel-Ghany, Salah E., author; Day, Irene, author; Heuberger, Adam L., author; Broeckling, Corey D., author; Reddy, Anireddy S. N., author; Elsevier Inc., publisher
    1,2,4-butanetriol (butanetriol) is a useful precursor for the synthesis of the energetic material butanetriol trinitrate and several pharmaceutical compounds. Bacterial synthesis of butanetriol from xylose or arabinose takes place in a pathway that requires four enzymes. To produce butanetriol in plants by expressing bacterial enzymes, we cloned native bacterial or codon optimized synthetic genes under different promoters into a binary vector and stably transformed Arabidopsis plants. Transgenic lines expressing introduced genes were analyzed for the production of butanetriol using gas chromatography coupled to mass spectrometry (GC-MS). Soil-grown transgenic plants expressing these genes produced up to 20 µg/g of butanetriol. To test if an exogenous supply of pentose sugar precursors would enhance the butanetriol level, transgenic plants were grown in a medium supplemented with either xylose or arabinose and the amount of butanetriol was quantified. Plants expressing synthetic genes in the arabinose pathway showed up to a forty-fold increase in butanetriol levels after arabinose was added to the medium. Transgenic plants expressing either bacterial or synthetic xylose pathways, or the arabinose pathway showed toxicity symptoms when xylose or arabinose was added to the medium, suggesting that a by-product in the pathway or butanetriol affected plant growth. Furthermore, the metabolite profile of plants expressing arabinose and xylose pathways was altered. Our results demonstrate that bacterial pathways that produce butanetriol can be engineered into plants to produce this chemical. This proof-of-concept study for phytoproduction of butanetriol paves the way to further manipulate metabolic pathways in plants to enhance the level of butanetriol production.
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    Primary and secondary metabolism in Centaurea maculosa and their potential roles in invasion biology
    (Colorado State University. Libraries, 2008) Broeckling, Corey D., author; Vivanco, Jorge M., advisor
    Centaurea maculosa is plant species native to Eurasia which has become invasive in North America, in part through allelopathic behavior. Allelopathy remains a highly debated subject, and a more firm understanding is necessary. C. maculosa is reported to secrete catechin as an allelochemical that is toxic to North American native plants. In this dissertation, a novel colorimetric assay for use in detection of catechin from soils is described and validated. This assay is highly sensitive, selective, and fast, which should allow for more detailed measures of catechin under field conditions, and ultimately a better understanding of the variation in catechin accumulation. To increase the depth of understanding of catechin biosynthesis, I report the cloning and characterization of C. maculosa dihydroflavonol reductase (CmDFR), a gene very likely to be necessary for catechin biosynthesis. To expand our view of metabolism beyond catechin, metabolome analysis is applied to field collected plant material, and demonstrate that the physiology of C. maculosa varies with the surrounding plant community -- plants growing in patches with high C. maculosa density tend to be accumulate higher levels of secondary metabolites than plants growing in the company of few conspecifics. Finally, the results of a study that clarifies the role of root exudates in structuring the soil fungal community are presented in the context of invasion biology.