Metabolic engineering of Vitamin B1 and Vitamin B6 in staple crops
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1 Research Collection Doctoral Thesis Metabolic engineering of Vitamin B1 and Vitamin B6 in staple crops Author(s): Li, Kuan-Te Publication Date: 2014 Permanent Link: Rights / License: In Copyright - Non-Commercial Use Permitted This page was generated automatically upon download from the ETH Zurich Research Collection. For more information please consult the Terms of use. ETH Library
2 DISS. ETH NO METABOLIC ENGINEERING OF VITAMIN B1 AND VITAMIN B6 IN STAPLE CROPS A thesis submitted to attain the degree of DOCTOR OF SCIENCES of ETH ZURICH (Dr. sc. ETH Zurich) presented by Kuan-Te Li M.Sc. in Agronomy, National Taiwan University (Taiwan) born on 3 rd August 1980 citizen of Taiwan accepted on the recommendation of Prof. Dr. Wilhelm Gruissem, examiner Prof. Dr. Teresa B. Fitzpatrick, co-examiner Dr. Hervé Vanderschuren, co-examiner 2014
3 Abstract Vitamin B1 and vitamin B6 are essential micronutrients for human health and plant growth. Both vitamins serve as cofactors in metabolic pathways, and are widely known as healthpromoting compounds. Plants use THI1 and THIC genes to produce thiazole and pyrimidine moieties separately, and subsequently couple these two moieties by a condensation to generate vitamin B1. The vitamin B6 de novo biosynthesis pathway involves only two genes, PDX1 and PDX2, in plants. Vitamin B1 and vitamin B6 deficiencies are associated with various chronic diseases including mental and physical disorders. The thesis aims at improving the vitamin B1 and vitamin B6 content in both cassava and rice via genetic engineering. Strategies to increase vitamin B6 content in cassava and rice were based on the constitutive or edible organ-specific expression of Arabidopsis PDX1.1 and PDX2 genes. Transgenic cassava showed a maximal 10-fold increase in vitamin B6 content in leaves using the constitutive expression strategy and a maximal 12-fold increase in storage roots using rootspecific expression strategy under field conditions. Transgenic rice showed up to 2-fold increase of vitamin B6 content in unpolished rice grains of both constitutive and endospermspecific expression vectors. The results demonstrate the potential of vitamin B6 biofortification in cassava and rice. Strategies to increase vitamin B1 content by constitutive expression of endogenous THI1 and/or THIC genes have been conducted in cassava and rice. Transgenic cassava displayed no increase or even decreased levels of vitamin B1 content in leaves and storage roots. Moreover, adverse effects on plant growth (e.g. chlorotic leaf, die back of top) were observed in several transgenic cassava lines. Transgenic rice showed no consistent increase of vitamin B1 content. Some individual transgenic rice plants over-expressing THI1 or THIC displayed higher vitamin B1 content at the seedling stage. Additionally, transgenic plants based on constitutive or edible organ-specific expression of Arabidopsis THIC were used to evaluate the impact of exogenous THIC on cassava and rice. Transgenic cassava using both constitutive and root-specific expression strategies showed decreased levels of vitamin B1 in leaves and storage roots of several lines. Transgenic rice showed up to 1.4-fold increase at seedling stage using the constitutive expression strategy and up to 1.6-fold increase in polished seeds using endosperm-specific expression strategy. The feasibility of vitamin B6 biofortification in cassava and rice was proven and is described in the thesis. It offers a sustainable and cost-effective approach to alleviate vitamin B6 deficiency in population having a diet mostly relying on cassava or rice consumption. Further characterization of transgenic plants expressing THI1 and THIC will help to investigate the vitamin B1 biosynthesis in cassava and rice. iii
4 Zusammenfassung Vitamin B1 und B6 sind essentielle Mikronährstoffe für die menschliche Gesundheit und das Pflanzenwachstum. Beide Vitamine dienen als Kofaktoren in Stoffwechselwegen und sind allgemein als gesundheitsfördernde Verbindungen bekannt. In Pflanzen werden Thiazol- und Pyrimidin-Einheiten separat mittels THI1 bzw. THIC hergestellt und anschließend zur Synthese von Vitamin B1 verbunden. In die pflanzliche de novo Biosynthese von Vitamin B6 sind nur die zwei Gene PDX1 und PDX2 involviert. Vitamin B1 und Vitamin B6 Mangel wird mit verschiedenen chronischen Krankheiten wie psychischen oder physischen Störungen in Verbindung gebracht. Das Ziel dieser Arbeit ist die Verbesserung des Vitamin B1 und Vitamin B6 Gehaltes in Cassava und Reis mittels gentechnologischen Methoden. Zur Steigerung des Vitamin B6 Gehaltes wurden PDX1.1 und PDX2 aus Arabidopsis konstitutiv oder Speiseorgan-spezifisch in Cassava und Reis exprimiert. Transgene Cassava zeigten unter Feldbedingungen eine maximal 10-fache Steigerung des Vitamin B6 Gehaltes in den Blättern mittels konstitutiver Expression und eine maximal 12-fache Erhöhung in Speicherwurzeln mittels Wurzel-spezifischer Expression. In unpolierten Körnern von konstitutiv und Endosperm-spezifisch PDX1.1/PDX2 exprimierenden Reispflanzen wurden bis zu 2-fach erhöhte Mengen an Vitamin B6 gemessen. Diese Ergebnisse demonstrieren das Potenzial von Vitamin B6 Biofortifikation in Cassava und Reis. Zur Steigerung des Vitamin B1 Gehaltes wurden die endogenen Gene THI1 und/oder THIC konstitutiv in Cassava und Reis exprimiert. In Blättern und Speicherwurzeln von transgenen Cassava wurde keine Zunahme aber teilweise eine Abnahme an Vitamin B1 beobachtet. Darüber hinaus zeigten sich nachteilige Auswirkungen auf das Pflanzenwachstum (z.b. chlorotische Blätter, Absterben der Spitze) in mehreren transgenen Cassava Linien. Transgene Reispflanzen zeigten keine konsistente Erhöhung des Vitamin B1 Gehaltes, nur im Keimlingsstadium wurden in einigen THI1 oder THIC überexprimierenden Pflanzen erhöhte Konzentrationen gemessen. Zusätzlich wurde THIC aus Arabidopsis konstitutiv und Speiseorgan-spezifisch in Maniok und Reis überexprimiert. Unabhängig von der Expressionsstrategie wurden in Blättern und Speicherwurzeln von transgenen Cassava verminderte Vitamin B1 Mengen gemessen. Transgene Reispflanzen zeigten unter konstitutiver Expression eine maximal 1,4-fache Steigerung des Vitamin B1 Gehaltes im Keimlingsstadium und bis zu 1,6-fache Erhöhung in polierten Körnern unter Endosperm-spezifischer Expression. Die Machbarkeit von Vitamin B6 Biofortifikation in Maniok und Reis wurden in dieser Arbeit nachgewiesen und beschrieben und bietet einen nachhaltigen und kostengünstigen Ansatz zur Linderung von Vitamin B6 Mangel in einer Bevölkerung, die zur Ernährung hauptsächlich iv
5 auf Reis und Cassava angewiesen ist. Die weitere Charakterisierung von THI1 und/oder THIC exprimierenden Pflanzen wird zum Verständnis der Vitamin B1 Biosynthese in Cassava und Reis beitragen. v
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Research Collection Doctoral Thesis To sense or not to sense viral RNA - essentials of coronavirus innate immune evasion Author(s): Kindler, Eveline Patricia Publication Date: 2015 Permanent Link: https://doi.org/10.3929/ethz-a-010560814
Grow rich and clean up later? Joint effects of international integration and democracy on environmental quality in developing countries
Research Collection Doctoral Thesis Grow rich and clean up later? Joint effects of international integration and democracy on environmental quality in developing countries Author(s): Ruoff, Gabriele Publication
Accountability and Internet of things applications
Research Collection Doctoral Thesis Accountability and Internet of things applications Author(s): Boos, Daniel Publication Date: 2011 Permanent Link: https://doi.org/10.3929/ethz-a-006687829 Rights / License:
Research Collection. Crystal structures and crystal chemistry of ternary LnT₂X type intermetallics. Doctoral Thesis. ETH Library
Research Collection Doctoral Thesis Crystal structures and crystal chemistry of ternary LnT₂X type intermetallics Author(s): Xue, Bingzhang Publication Date: 1994 Permanent Link: https://doi.org/10.3929/ethz-a-000961977
Schlafstörungen durch nächtlichen Alltagslärm Aus der Public Health Perspektive
Research Collection Doctoral Thesis Schlafstörungen durch nächtlichen Alltagslärm Aus der Public Health Perspektive Author(s): Omlin, Sarah Publication Date: 2011 Permanent Link: https://doi.org/10.3929/ethz-a-007316362
Research Collection. Constraint-based surface processing for geometric modeling and architecture. Doctoral Thesis. ETH Library
Research Collection Doctoral Thesis Constraint-based surface processing for geometric modeling and architecture Author(s): Eigensatz, Michael Publication Date: 2010 Permanent Link: https://doi.org/10.3929/ethz-a-006699489
Research Collection. Computational modeling of tumor growth. Doctoral Thesis. ETH Library. Author(s): Lloyd, Bryn Alexander. Publication Date: 2010
Research Collection Doctoral Thesis Computational modeling of tumor growth Author(s): Lloyd, Bryn Alexander Publication Date: 2010 Permanent Link: https://doi.org/10.3929/ethz-a-006088514 Rights / License:
Research Collection. Working Paper. ETH Library. Author(s): Bächi, Beat. Publication Date: 2001
Research Collection Working Paper Kommunikationstechnologischer und sozialer Wandel. "Der schweizerische Weg zur digitalen Kommunikation" (1960-1985) "der schweizerische Weg zur digitalen Kommunikation"
Integer Convex Minimization in Low Dimensions
DISS. ETH NO. 22288 Integer Convex Minimization in Low Dimensions A thesis submitted to attain the degree of DOCTOR OF SCIENCES of ETH ZURICH (Dr. sc. ETH Zurich) presented by TIMM OERTEL Diplom-Mathematiker,
Research Collection. Doctoral Thesis. ETH Library. Author(s): Dinkel, Martin. Publication Date: 2011
Research Collection Doctoral Thesis Regulation of the NoCut Checkpoint and its Interplay with the Abscission Pathway Prevents Premature Fission of the Plasma Membrane in Saccharomyces Cerevisiae Author(s):
Biometric authentication system using human gait
Research Collection Doctoral Thesis Biometric authentication system using human gait Author(s): Cattin, Philippe Publication Date: 2002 Permanent Link: https://doi.org/10.3929/ethz-a-004407948 Rights /
Research Collection. Promoter analysis and transcriptional regulation of the CMT1Adisease gene peripheral myelin protein PMP22.
Research Collection Doctoral Thesis Promoter analysis and transcriptional regulation of the CMT1Adisease gene peripheral myelin protein PMP22 Author(s): Maier, Marcel Publication Date: 2003 Permanent Link:
Analysis, optimization, and synthesis of planar integrated lightwave circuits for WDM applications
Research Collection Doctoral Thesis Analysis, optimization, and synthesis of planar integrated lightwave circuits for WDM applications Author(s): Spühler, Michael-Martin Publication Date: 2000 Permanent
Engineering of synthetic mammalian gene networks and their cross-kingdom conservation
Research Collection Doctoral Thesis Engineering of synthetic mammalian gene networks and their cross-kingdom conservation Author(s): Gitzinger, Marc Publication Date: 2010 Permanent Link: https://doi.org/10.3929/ethz-a-006425023
On the List Update Problem
DISS. ETH No. 14529, 2002 On the List Update Problem A dissertation submitted to the Swiss Federal Institute of Technology, ETH Zürich for the degree of Doctor of Technical Sciences presented by Christoph
Research Collection. Improving geometric calibration methods for multi-axis machining centers by examining error interdependencies effects
Research Collection Doctoral Thesis Improving geometric calibration methods for multi-axis machining centers by examining error interdependencies effects Author(s): Bringmann, Bernhard Publication Date:
Informationstechnische Unterstützung der frühen Phasen der Produkt-Innovation
Research Collection Doctoral Thesis Informationstechnische Unterstützung der frühen Phasen der Produkt-Innovation Author(s): Nyffenegger, Felix Publication Date: 2006 Permanent Link: https://doi.org/10.3929/ethz-a-005348615
The measurement of the oxygenation status of the newborn infant
Research Collection Doctoral Thesis The measurement of the oxygenation status of the newborn infant Author(s): Wolf, Martin Peter Publication Date: 1996 Permanent Link: https://doi.org/10.3929/ethz-a-001630372
Ueber den Einfluss von Salz-Zusätzen auf das Phasengleichgewicht wässeriger Ameisensäure
Research Collection Doctoral Thesis Ueber den Einfluss von Salz-Zusätzen auf das Phasengleichgewicht wässeriger Ameisensäure Author(s): Johnsen, Bjarne Karth Publication Date: 1954 Permanent Link: https://doi.org/10.3929/ethz-a-000089716