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1

Jin, Xin. "Isoprenoid and flavonoid biosynthesis and regulation in higher plants." Doctoral thesis, Universitat de Lleida, 2019. http://hdl.handle.net/10803/667579.

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Aquesta tesi es centra principalment en l'anàlisi funcional i la caracterització dels gens que codifiquen per a alguns metabòlits secundaris i en l’estudi de la seva regulació en les plantes. Els objectius generals varen ser (a) entendre millor la regulació transcripcional del gen de la biosíntesi dels carotenoids, la β-carotè hidroxilasa 2 (BCH2) en el blat de moro i (b) l'anàlisi funcional de les dues isopentenil difosfat isomerasas (OsIPPI) d'arròs i determinar la seva localització subcel·lular. Simultàniament, es va estudiar com la llum afecta la via metabòlica i a la producció de pelargonidina en l'arròs; es van identificar també els gens essencials de la seva biosíntesi en Gentiana lutea L. var. aurantiaca. Les plantes de blat de moro i arròs es varen transformar amb els gens dels factors de transcripció ZmMYB i ZmPBF. Es va analitzar l’expressió gènica transitòria i es va realitzar transformació estable. Els resultats obtinguts indiquen que tant ZmPBF com ZmGAMYB poden transactivar l'expressió de ZmBCH2 a l’endosperm del blat de moro, i ZmPBF i ZmGAMYB transactiven independentment el promotor de ZmBCH2 en arròs. Els dos paràlegs de IPPI (OsIPPI1 i OsIPPI2) aïllats prèviament en arròs varen tenir un patró d'expressió diferent; l'ARNm de OsIPPI1 va ser més abundant que l'ARNm de OsIPPI2 en tots els teixits. Es va usar la microscòpia de fluorescència confocal i microscòpia inmunoelectrónica per determinar la localització de les dues proteïnes. Aquestes es localitzen en el reticle endoplasmàtic (RE), així com en els peroxisomes i les mitocòndries, mentre que només es va detectar OsIPPI2 en els plastidis. La detecció d'ambdues isoformes en el RE indica que DMAPP es pot sintetitzar de novo en aquest compartiment. Diferents tècniques com UPLC, GC-MS i qRT-PCR també es varen utilitzar per perfilar els metabòlits primaris i secundaris i l'expressió gènica relacionada en plàntules d'arròs des-etioladas. Els resultats varen revelar que el metabolisme primari i secundari i els gens corresponents estan regulats per la llum, especialment en la biosíntesi d'isoprenoides en fulles d'arròs. Onze derivats de pelargonidina es varen identificar en els pètals de G. lutea i es varen perfilar els gens de la seva via de biosíntesi, revelant que DFR, ANS i 3GT afecten principalment a l'acumulació dels glucòsids de pelargonidina. Tots aquests resultats suggereixen la idea que la biosíntesi dels carotenoids en plantes superiors és regulada a diferents nivells.
Esta tesis se centra principalmente en el análisis funcional y en la caracterización de los genes que codifican para algunos metabolitos secundarios y en el estudio de su regulación en las plantas. Los objetivos generales fueron (a) profundizar en el conocimiento de la regulación transcripcional del gen de la biosíntesis de los carotenoides, la β-caroteno hidroxilasa 2 (BCH2) en el maíz, y (b) analizar la función de las dos isopentenil difosfato isomerasas (OsIPPI) de arroz, determinando además su localización subcelular. Simultáneamente, se estudió cómo la luz afecta a la vía metabólica y a la producción de pelargonidina en el arroz; se identificaron también los genes esenciales de su biosíntesis en Gentiana lutea L. var. aurantiaca. Las plantas de maíz y arroz se transformaron con los genes de los factores de transcripción ZmMYB y ZmPBF. Se analizó la expresión génica transitoria y se realizó transformación estable. Los resultados obtenidos indicaron que tanto ZmPBF como ZmGAMYB pueden transactivar la expresión de ZmBCH2 en endospermo de maíz, y ZmPBF y ZmGAMYB transactivar independientemente el promotor de ZmBCH2 en arroz. Los dos parálogos de IPPI (OsIPPI1 y OsIPPI2) aislados previamente en arroz tuvieron un patrón de expresión diferente; el ARNm de OsIPPI1 fue más abundante que el ARNm de OsIPPI2 en todos los tejidos. Se usó la microscopía de fluorescencia confocal y microscopía inmunoelectrónica para determinar la localización de ambas proteínas. Estas se localizan en el retículo endoplásmico (RE), así como en los peroxisomas y las mitocondrias, mientras que solo se detectó OsIPPI2 en los plastidios. La detección de ambas isoformas en el RE indica que DMAPP se puede sintetizar de novo en este compartimiento. Diferentes técnicas como UPLC, GC-MS y qRT-PCR también se utilizaron para perfilar los metabolitos primarios y secundarios y la expresión génica en plántulas de arroz des-etioladas. Los resultados revelaron que los genes involucrados en la en el metabolismo primario y secundario están regulados por la luz, especialmente en la biosíntesis de isoprenoides en hojas de arroz. Once derivados de pelargonidina se identificaron en los pétalos de G. lutea y se perfilaron los genes de la vía de biosíntesis, revelando que DFR, ANS y 3GT afectan principalmente a la acumulación de los glucósidos de pelargonidina. Todos estos resultados contribuyen al conocimiento, a diferentes niveles, de la regulación de las rutas biosinteticas de los carotenoides en plantas superiores.
This thesis mainly focuses on functional analysis and characterization of a number of secondary metabolite biosynthetic genes and the regulation of the corresponding secondary metabolite biosynthetic pathway in plants. The overall aims were to elucidate the transcriptional regulation of β-carotene hydroxylase 2 gene (BCH2) in maize, the functional analysis of rice isopentenyl diphosphate isomerases (OsIPPI), and determine their subcellular localization. Simultaneously, the influence of light on the metabolic pathway in rice was studied and the pelargonidin quantification and essential pelargonidin biosynthesis genes in Gentiana lutea L. var. aurantiaca were identified. Maize and rice plants were transformed with transcription factor genes ZmMYB and ZmPBF, via transient gene expression and stable transformation respectively. The results indicated that both ZmPBF and ZmGAMYB can transactivate ZmBCH2 expression in maize endosperm and ZmPBF and ZmGAMYB independently transactivate the ZmBCH2 promoter in rice. The two IPPI paralogs (OsIPPI1 and OsIPPI2) isolated previously in rice had a different expression pattern; OsIPPI1 mRNA was more abundant than OsIPPI2 mRNA in all tissues. Confocal fluorescence microscopy and immuno-electron microscopy were used to determine the localization of both proteins. These localized to the endoplasmic reticulum (ER) as well as peroxisomes and mitochondria, whereas only OsIPPI2 was detected in plastids. The detection of both isoforms in the ER indicates that DMAPP can be synthesized de novo in this compartment. UPLC, GC-MS and qRT-PCR were used to profile the primary and secondary metabolites and gene expression in de-etioleted rice seedlings. The results revealed both primary and secondary metabolism and the corresponding genes are regulated by light, especially isoprenoids biosynthesis in rice leaves. Eleven pelargonidin derivatives were identified in the petals of G. lutea and the biosynthetic pathway genes were profiled, revealing DFR, ANS and 3GT mainly affect the accumulation of pelargonidin glucosides. Collectively my results provide novel insights of the regulation of carotenoid and flavonoid biosynthesis in higher plants at different levels.
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2

Savill, Julia. "Carotenoid biosynthesis in higher plants." Thesis, Liverpool John Moores University, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.313103.

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3

Milborrow, Barry Vaughan Biotechnology &amp Biomolecular Sciences Faculty of Science UNSW. "Biosynthesis of abscisic acid in plants." Awarded by:University of New South Wales. Biotechnology & Biomolecular Sciences, 2007. http://handle.unsw.edu.au/1959.4/42883.

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4

Parry, Andrew David. "Abscisic acid biosynthesis in higher plants." Thesis, Cardiff University, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.328480.

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5

Macaulay, Keith Malcolm. "Salicylic acid biosynthesis in higher plants." Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609202.

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6

Hu, Chien-an Andy. "Osmoregulation and proline biosynthesis in plants /." The Ohio State University, 1993. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487843688956923.

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7

Wheeler, Glen L. "The biosynthesis of ascorbic acid in plants." Thesis, University of Exeter, 2000. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.531672.

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8

Ashurst, Jennifer Lilian. "Investigation of pantothenate biosynthesis in higher plants." Thesis, University of Cambridge, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.621651.

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9

Okunishi, Tomoya. "Stereochemistry of Lignan Biosynthesis in Thymelaeaceae Plants." Kyoto University, 2003. http://hdl.handle.net/2433/149001.

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Kyoto University (京都大学)
0048
新制・課程博士
博士(農学)
甲第10276号
農博第1348号
新制||農||869(附属図書館)
学位論文||H15||N3797(農学部図書室)
UT51-2003-H697
京都大学大学院農学研究科応用生命科学専攻
(主査)教授 島田 幹夫, 教授 桑原 保正, 教授 坂田 完三
学位規則第4条第1項該当
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10

Hamilton, John T. G. "Biosynthesis of organofluorine compounds in plants and bacteria." Thesis, Queen's University Belfast, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.388092.

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11

Chakauya, Ereck. "Effect of manipulating pantothenate biosynthesis in higher plants." Thesis, University of Cambridge, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.614866.

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12

Gutiérrez, García Laura. "Unveiling the biological role of stigmasterol biosynthesis in tomato plants." Doctoral thesis, Universitat de Barcelona, 2019. http://hdl.handle.net/10803/667925.

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In plants, the sterol biosynthetic pathway leads to the production of a complex mixture of end-products, among which β-sitosterol, stigmasterol, campesterol, and cholesterol are the most abundant ones. Stigmasterol, the end-product of the 24-ethyl sterol branch is synthesized from β-sitosterol by the action of sterol C22-desaturase (C22DES). Despite C22DES was identified about a decade ago, there is still a lack of knowledge about several relevant aspects related to the structure and function of this enzyme. Furthermore, several studies have reported the occurrence of changes in the stigmasterol to β-sitosterol ratio (STIG/SIT) during plant development and in their responses to environmental stimuli which have been related with the activity of C22DES. However, the biological relevance of the changes in stigmasterol levels is currently unknown. Based on this, the main objective of present work has been the elucidation of the role of stigmasterol during plant growth and development as well as in its response to environmental challenges. Tomato (Solanum lycopersicum cv. MicroTom) was selected for this studies taking advantage that is one of the few plants in which C22DES is encoded by a single-copy gene. The goals of this thesis have been: (1) the functional and structural characterization of C22DES in terms of subcellular localization and mechanism of action, and (2) the evaluation of the role of stigmasterol biosynthesis in tomato plant growth, development and in responses to biotic and abiotic stresses. The results obtained in this thesis have confirmed that C22DES is targeted and retained in the endoplasmic reticulum (ER)-membrane. C22DES consists of two well-differentiated domains: a single N-terminal transmembrane-helix domain (TMH1), which is sufficient for the ER-membrane targeting and retention, and a globular domain that also contacts with the ER-membrane. The globular domain may also interact and be retained in the ER in the absence of TMH1, but it is enzymatically inactive, revealing the requirement of the N-terminal membrane domain for enzyme activity. The in silico analysis of the TMH1 region revealed several features that could be involved in substrate recognition and binding. Transgenic tomato plants with altered STIG/SIT ratios have been generated by overexpression, amiRNA-mediated silencing, and knock-out of the C22DES gene. The obtained results have shown that lack of stigmasterol is not lethal for the plant since the C22des- knock-out plants are able to complete their life cycle. However, C22des- knock- out mutant showed several phenotypic abnormalities related to growth and developmental processes. The C22des- knock-out plants also show increased susceptibility to Botrytis cinerea infection. The data provided in this thesis contribute to expand the current knowledge on the mechanism of action of stigmasterol and C22DES in tomato plants and set the basis for further studies focused on unraveling the mechanisms involved in the regulation of C22DES activity. The C22des- knock-out mutant generated in this work also provides a unique and highly relevant tool to evaluate the role of stigmasterol in plants.
En plantas, la ruta de biosíntesis de los esteroles da lugar a la generación de una compleja mezcla de productos finales, entre los cuales, el β-sitosterol, estigmasterol, campesterol, colesterol son los más abundantes. El estigmasterol es el producto final en la rama de los 24-etil esteroles, y se sintetiza a partir del β-sitosterol por la acción de la enzima esterol C22-desaturasa (C22DES). A pesar de que C22DES fue identificada hace una década, aún hace falta mayor conocimiento acerca de algunos aspectos relevantes relacionados con la estructura y función de esta enzima. Además, algunos estudios han descrito la existencia de cambios en la relación estigmasterol a β-sitosterol (STIG/SIT) durante el desarrollo de la planta y su respuesta a estímulos ambientales, que se han relacionado con la actividad de C22DES. Sin embargo, aún se desconoce la relevancia biológica de los cambios en los niveles de estigmasterol. Basándonos en esto, el objetivo principal del presente trabajo ha sido el esclarecimiento del papel del estigmasterol durante el crecimiento de la planta y el desarrollo, así como en su respuesta a desafíos ambientales. Para este estudio se seleccionó el tomate (Solanum lycopersicum) como planta modelo aprovechando que es una de las pocas especies vegetales en las que C22DES está codificada por un gen de copia única. Los objetivos de esta tesis han sido: (1) la caracterización funcional y estructural de C22DES en términos de localización subcelular y mecanismo de acción, y (2) la evaluación del papel de la biosíntesis de estigmasterol sobre el crecimiento, desarrollo y respuestas a estrés biótico y abiótico en plantas de tomate. Los resultados obtenidos en esta tesis han confirmado que C22DES se dirige y retiene en la membrana del retículo endoplásmico (RE). C22DES consiste en dos dominios bien diferenciados: un dominio hélice transmembrana en el extremo amino-terminal (TMH1), que es suficiente para su direccionamiento y retención en la membrana del RE, y un dominio globular que también mantiene zonas de contacto con la membrana del RE. El dominio globular también puede interaccionar y ser retenido en el RE en ausencia de TMH1, pero es enzimáticamente inactivo, lo que revela la necesidad del dominio transmembrana amino-terminal para la actividad enzimática. Los análisis in silico de la región TMH1 revelaron algunas características que pueden estar implicadas en el reconocimiento y unión de sustrato. Se generaron plantas transgénicas de tomate con relaciones STIG/SIT alteradas mediante la sobreexpresión, silenciamiento mediado por amiRNA, y la eliminación del gen C22DES. Los resultados obtenidos mostraron que la ausencia de estigmasterol no es letal para la planta, ya que las plantas del genotipo mutante nulo C22des- son capaces de completar su ciclo de vida. Sin embargo, este mutante mostró algunas anormalidades fenotípicas relacionadas con procesos de crecimiento y desarrollo. Las plantas mutantes C22des- también mostraron aumentada su susceptibilidad a la infección por Botrytis cinerea. Los datos proporcionados en esta tesis contribuyen a aumentar el conocimiento actual sobre los mecanismos de acción del estigmasterol y C22DES en plantas de tomate y sienta las bases para futuros estudios dedicados a desentrañar los mecanismos involucrados en la regulación de la actividad C22DES. El mutante C22des- generado en este trabajo también proporciona una herramienta única y de gran importancia para evaluar el papel del estigmasterol en las plantas.
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13

Zhang, Dong-Xiu. "Loline alkaloid biosynthesis gene expression in epichloë endophytes of grasses." Lexington, Ky. : [University of Kentucky Libraries], 2008. http://hdl.handle.net/10225/785.

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Thesis (Ph. D.)--University of Kentucky, 2008.
Title from document title page (viewed on May 12, 2008). Document formatted into pages; contains: xvi, 221 p. : ill. (some col.). Includes abstract and vita. Includes bibliographical references (p. 214-219).
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14

Gross, Jeferson. "The biosynthesis of phylloquinone(vitamin K1) in higher plants." Diss., lmu, 2006. http://nbn-resolving.de/urn:nbn:de:bvb:19-63535.

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15

Lundmark, Åstot Crister. "Cytokinins in higher plants : biosynthesis and interaction with auxins /." Umeå : Swedish Univ. of Agricultural Sciences (Sveriges lantbruksuniv.), 2000. http://epsilon.slu.se/avh/2000/91-576-5866-8.pdf.

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16

Rizk, Sandra E. "Biosynthesis and assembly of pectin and glucuronoarabinoxylan in plants." Thesis, University of Glasgow, 2000. http://theses.gla.ac.uk/5655/.

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Nascent pectin and glucuronarabinoxylan (GAX), synthesized in vitro by membrane-bound enzymes from etiolated pea (Pisum sativum L.) epicotyls, were found to bind to pea xyloglucan in a pH-dependent manner. The binding was maximum at low pH (3-4), and decreased to almost zero at pH 6. The binding seemed to occur instantaneously, to be non-covalent, and to require both terminal fucose residues of xyloglucan, in addition to the non-reduced acid residues of GAX and pectin. Removal by protease of the proteins attached to nascent pectin and GAX, greatly reduced the maximum binding and abolished the pH-dependence. The proteins involved seem to have approximate molecular weights of 14 and 94 kDa. The pH-dependent binding of nascent pectin and GAX is not completely specific to xyloglucan, since some binding occurred to a range of other matrix polysaccharides, though at a lower level than to pea xyloglucan. Newly-deposited pectin was extracted from peas that were incubated with radioactively labelled sucrose. It was shown to behave in a similar manner as nascent pectin, exhibiting the same pH-dependent binding pattern to xyloglucan. Protease treatment of pectin decreased the binding, indicating the possible presence of proteins attached to pectin in the cell wall, and revealing the role of those proteins in the interaction of pectin with other matrix polysaccharides, particularly xyloglucan. The pH-dependence of the binding suggests a functional interaction with the mechanisms that control growth, since the wall pH decreases when elongation growth is initiated. The proteins involved would play a significant role in cell-wall assembly and cell-wall elongation.
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17

Harris, Darby M. "MOLECULAR AND CHEMICAL DISSECTION OF CELLULOSE BIOSYNTHESIS IN PLANTS." UKnowledge, 2011. http://uknowledge.uky.edu/pss_etds/3.

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Plant cell walls are complex structures that must not only constrain cellular turgor pressure but also allow for structural modification during the dynamic processes of cell division and anisotropic expansion. Cell walls are composed of highly glycosylated proteins and polysaccharides, including pectin, hemicellulose and cellulose. The primary cell wall polysaccharide is cellulose, a polymer composed of high molecular weight !- 1,4-glucan chains. Although cellulose is the most abundant biopolymer on Earth, there is still a lot to learn about its biosynthesis and regulation. This research began by applying a variety of analytical techniques in an attempt to understand differences in cell wall composition and cellulose structure within the plant body, between different plant species and as a result of acclimation by the plant to different environmental conditions. Next, a number of different Arabidopsis thaliana lines possessing mutations affecting cell wall biosynthesis were analyzed for changes in cellulose structure (crystallinity) and biomass saccharification efficiency. One of these mutants, isoxaben resistance1-2 (ixr1- 2), which contains a point mutation in the C-terminal transmembrane region (TMR) of cellulose synthase 3 (CESA3), exhibited a 34% lower biomass crystallinity index and a 151% improvement in saccharification efficiency relative to that of wild-type. The culmination of this research began with a chemical screen that identified the molecule quinoxyphen as a primary cell wall cellulose biosynthesis inhibitor. By forward genetics, a semi-dominant mutant showing strong resistance to quinoxyphen named aegeus was identified in A. thaliana and the resistance locus mapped to a point mutation in the TMR of CESA1. cesa1aegeus occurs in a similar location to that of cesa3ixr1-2, illustrating both subunit specificity and commonality of resistance locus. These drug resistant CESA TMR mutants are dwarfed and have aberrant cellulose deposition. High-resolution synchrotron X-ray diffraction and 13C solid-state nuclear magnetic resonance spectroscopy analysis of cellulose produced from cesa1aegeus, cesa3ixr1-2 and the double mutant shows a reduction in cellulose microfibril width and an increase in mobility of the interior glucan chains of the cellulose microfibril relative to wild-type. These data demonstrate the importance of the TMR region of CESA1 and CESA3 for the arrangement of glucan chains into a crystalline cellulose microfibril in primary cell walls.
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18

Horstmann, Carl Ulrich. "Manipulating cell wall biosynthesis in yeast and higher plants." Thesis, Stellenbosch : University of Stellenbosch, 2010. http://hdl.handle.net/10019.1/5288.

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Thesis (MSc (Genetics))--University of Stellenbosch, 2010.
Includes bibliography.
Title page: Dept. of Genetics, Faculty of Science.
ENGLISH ABSTRACT: Undeniably, changes in the environment and dwindling traditional energy resources have resulted in the search for viable, renewable energy alternatives such as biofuels. Cellulose is one of the most abundant polymers on earth and can be converted to simple sugars and fermented to ethanol biofuel fairly easily. Cellulose rich biomass that can serve to supply ethanol biofuel production can be sourced from unexploited agricultural waste. The main drawback to using vegetative tissue as opposed to harvested food stocks from crops results from the structural properties of plant cell walls. Although cellulose is abundant, the contaminating hemicellulose and lignin fibres within the cell wall matrix have a negative impact on the digestibility of the cellulose present. Thus, an important step in creating an effective biofuel production system from agricultural excess is developing crops with improved cell wall polymer characteristics that can be converted to ethanol more efficiently. This project consisted of two parts. Firstly, the aim was to assess lignin production in transgenic sugarcane transformed with a construct aimed at down-regulating the 4- (hydroxyl) cinnamoyl CoA ligase (4CL) gene in the lignin biosynthesis pathway. The second part of the project revolved around discovering the mechanism of impared cell growth caused by expressing the gene encoding cellulose synthase from a marine invertebrate, Ciona savignyi, in the yeast Saccharomyces cerevisiae. Several sugarcane lines that had been previously transformed with a hairpin RNAi construct aimed at down-regulating the 4CL gene in the monolignol biosynthesis pathway were subjected to analysis to determine if lignification had been reduced. Although the presence of the hairpin construct in the genomic DNA had been confirmed for all of the transgenic lines, there was no significant decrease in the lignin levels in any of the transgenic lines. PCR analysis of the mRNA and enzyme assays also confirmed that the 4CL gene was still being expressed. Ongoing work will determine the cause of the unsuccessful down-regulation. Previously, it had been proven that the cellulose synthase gene from C. savignyi could be functionally expressed in S. cerevisiae. However, cellulose production resulted in extremely retarded growth of colonies and cultures, to the point of the apparent death of the cultures. The aim of this part of the project was to determine the mechanism (either metabolic or physical) that causes this effect. To generate enough cell mass to perform metabolic analysis, several strategies to impede cellulose production in transgenic yeast were explored. Attempts to stop cellulose production and induce better growth by introducing Isoxaben (a traditional weed killer that targets cellulose synthases) into the growth medium used for the transgenic yeast proved unsuccessful. To control the expression of the transgene, it was attempted to clone the cellulose synthase gene into an expression system containing an inducible promoter. The cloning exercise proved extremely difficult and multiple attempts with several strategies proved unsuccessful. This process is still ongoing as the growth retarding process induced by cellulose production in yeast remains to be identified.
AFRIKAAANSE OPSOMMING: Dit is onontkenbaar dat veranderinge in die omgewing en minderwordende tradisionele energiebronne veroorsaak dat lewensvatbare en hernubare energiebronne soos biobrandstof gevind moet word. Sellulose is een van die mees volop polimere op aarde en kan redelik maklik omgeskakel word na eenvoudige suikers en gefermenteer word tot etanol-biobrandstof. Sellulose-ryk biomassa wat etanol-biobrandstof kan verskaf, kan herwin word van tot op hede ongebruikte landbou-afval. Die komplekse struktuur van plantselwande is die hoofstruikelblok in die omskakeling van vegetatiewe weefsel tot biobrandstof. Hoewel sellulose volop is, het die kontaminerende hemisellulose- en lignienvesels binne die selwand-matriks ’n negatiewe impak op die verteerbaarheid van die sellulose teenwoordig in die selwand. Daarom is ’n belangrike stap in die ontwikkeling van effektiewe biobrandstof-produksiesisteme vanaf landbou-afval om gewasse te ontwikkel met verbeterde selwandpolimeer-eienskappe wat etanol-produksie kan vergemakilik. Hierdie projek het bestaan uit twee dele. Eerstens was die doel om vas te stel of die lignienproduksie geaffekteer is in transgeniese suikerriet getransformeer met ’n konstruk wat mik om die 4-(hidroksie)-cinnamoyl CoA ligase (4CL) geen te af-reguleer in die lignienbiosintese- padweg. Die tweede deel van die projek het daarop gefokus om die meganisme te ondek wat die belemmerde selgroei veroorsaak, as gevolg van die uitdrukking van die geen wat kodeer vir sellulose-sintase in ’n mariene ongewerwelde, Ciona savignyi, in Saccharomyces cerevisiae. Verskeie suikerriet-lyne, wat voorheen getransformeer is met ’n haarnaald-RNAi-konstruk om die 4CL-geen te af-reguleer in die monolignol-biosintese-padweg, is onderwerp aan analise om vas te stel of lignifikasie verminder is. Hoewel die teenwoordigheid van die haarnaald-konstruk in die genomiese DNA bevestig is vir al die transgeniese lyne, was daar geen beduidende vermindering in die lignienvlakke in die transgeniese lyne nie. PKRanalise van die mRNA en ensiem-aktiwiteitstoetse het ook bevestig dat die 4CL-geen steeds uitgedruk word. Verdere ondersoek sal kan vasstel wat die oorsaak van die onsuksesvolle af-regulering is. Voorheen is bewys dat die sellulose-sintase-geen van C. savignyi funksioneel uitgedruk kon word in Saccharomyces cerevisiae. Egter, selluloseproduksie het die gevolg gehad dat groei in die transgeniese kolonies en kulture erg gestrem is, tot die punt dat die kulture dood voorgekom het. Die doel van hierdie deel van die projek was om vas te stel wat die meganisme (òf metabolies òf fisies) is wat hierdie verskynsel veroorsaak het. Om genoeg selmassa te genereer om metaboliese analise uit te voer, is verskeie strategieë om selluloseproduksie in transgeniese gis te verhinder, ondersoek. Pogings om selluloseproduksie te stop en om groei te verbeter deur Isoxaben by te voeg in die groeimedium gebruik vir transgeniese gis, was onsuksesvol. Isoxaben is ’n tradisionele onkruiddoder wat sellulose-sintases teiken en inhibeer. Om die uitdrukking van die transgeen te beheer, is ’n poging aangewend om dié sellulose-sintase-geen in ’n uitdrukking-sisteem te kloon met ’n induseerbare promotor. Die kloneringsoefening was uiters moeilik en veelvoudige pogings met verskeie strategieë was onsuksesvol. Hierdie proses moet verder gevoer word aangesien die groeistremmingsmeganisme veroorsaak deur selluloseproduksie in gis nog geïdentifiseer moet word.
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19

Terry, Christian James. "Gene expression and ABA biosynthesis in water stressed plants." Thesis, University of Nottingham, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308310.

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Tunc, Meral. "The molecular genetic regulation of thiamin biosynthesis in plants." abstract and full text PDF (free order & download UNR users only), 2008. http://0-gateway.proquest.com.innopac.library.unr.edu/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3307578.

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Roosens, Nancy. "Proline biosynthesis related to salt stress in higher plants." Doctoral thesis, Universite Libre de Bruxelles, 1999. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/211926.

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Addlesee, Hugh Alistair. "The terminal stages of chlorophyll biosynthesis in bacteria and plants." Thesis, University of Sheffield, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.265151.

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Runguphan, Weerawat. "Reprogramming alkaloid biosynthesis in Catharanthus roseus : synthetic biology in plants." Thesis, Massachusetts Institute of Technology, 2011. http://hdl.handle.net/1721.1/65274.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2011.
Vita. Cataloged from PDF version of thesis.
Includes bibliographical references.
The medicinal plant Madagascar periwinkle (Catharanthus roseus) produces over 130 monoterpene indole alkaloid (MIA) natural products. Many of these compounds have pharmaceutical value, such as the anticancer agents vinblastine and vincristine. Unnatural modifications can impart novel bioactivity to the parent natural product. Advances in synthetic biology and microbial engineering have allowed overproduction of natural products and their analogs in non-native organisms such as yeast and E. coli. However, re-engineering of plant pathways to yield "novel" products has been limited, particularly when compared to the successes achieved in prokaryotic systems. This thesis describes several strategies to re-engineer MIA biosynthesis in periwinkle to produce novel alkaloids. The first strategy involves the introduction of a biosynthetic enzyme with redesigned substrate specificity into periwinkle. The resulting transgenic plant culture produces a variety of unnatural alkaloid compounds when co-cultured with precursors that the re-engineered enzyme has been designed to accept. The second strategy improves upon this work by enabling periwinkle to autonomously synthesize precursor analogs in situ. Specifically, the prokaryotic halogenation machinery was introduced into the genome of periwinkle, which lacks the biosynthetic ability to produce halogenated compounds. These halogenases function within the context of the plant cell to generate halogenated precursor, which is then shuttled into MIA metabolism to yield halogenated alkaloids. Altogether, a new functional group-an organohalide-was introduced into plant secondary metabolism in a regioselective and predictable manner. The third strategy involves RNAi-mediated suppression of MIA biosynthesis in periwinkle. Alkaloid production was obliterated in the resulting transgenic plant culture. The silenced plant culture produces a variety of fluorinated alkaloids when co-cultured with fluorinated starting substrate. The yields of some unnatural alkaloids were improved since the natural precursor was absent. Finally, the fourth strategy describes chemical functionalization of halogenated MIAs. Postbiosynthetic chemical derivatizations of halogenated MIAs using palladium-catalyzed Suzuki-Miyaura cross-coupling reactions robustly afforded aryl and heteroaryl analogs of MIAs. Altogether, the work described in this thesis demonstrates the versatility of medicinal plants in the generation of unnatural alkaloids. Thus, despite their genetic complexity, plants are a viable platform for synthetic biology efforts.
by Weerawat Runguphan.
Ph.D.
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Zhang, Chun-sheng. "Regulation of proline biosynthesis in plants subjected to osmotic stress /." The Ohio State University, 1997. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487948807588165.

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Ahmed, Sheaza. "Metabolic Engineering of Plants by Manipulating Polyamine Transport and Biosynthesis." Bowling Green State University / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1507393675673047.

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Trojanowska, Miranda R. "The biosynthesis of saponins and phytosterols in oat roots." Thesis, University of Hull, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.310222.

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Al-Shakarchi, E. M. D. "Transformation of sterols in plant tissue cultures." Thesis, Bucks New University, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.384187.

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Shih, Chun-hat, and 施振翮. "Molecular characterization and metabolic engineering of flavonoid biosynthesis in higher plants." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2009. http://hub.hku.hk/bib/B41633829.

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Shih, Chun-hat. "Molecular characterization and metabolic engineering of flavonoid biosynthesis in higher plants." Click to view the E-thesis via HKUTO, 2009. http://sunzi.lib.hku.hk/hkuto/record/B41633829.

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Lehner, Bryan W. "Aggregation Pheromone Biosynthesis and Engineering in Plants for Stinkbug Pest Management." Thesis, Virginia Tech, 2019. http://hdl.handle.net/10919/100605.

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Stinkbugs (Pentatomidae) and other agricultural pests such as bark beetles and flea beetles are known to synthesize terpenoids as aggregation pheromones. Knowledge of the genes and enzymes involved in pheromone biosynthesis may allow engineering of pheromone biosynthetic pathways in plants to develop new forms of trap crops and agricultural practices for pest management. The harlequin bug, Murgantia histrionica, a specialist pest on crucifer crops, produces the sesquiterpene, murgantiol, as a male-specific aggregation pheromone. Similarly, the southern green stink bug, Nezara viridula, a generalist pest worldwide on soybean and other crops, releases sesquiterpene cis-/trans-(Z)-α-bisabolene epoxides as male-specific aggregation pheromone. In both species, enzymes called terpene synthases (TPSs) synthesize precursors of the aggregation pheromones, which are sesquipiperitol and (Z)-α-bisabolene as the precursor of murgantiol and cis-/trans-(Z)-α-bisabolene epoxide, respectively. We hypothesized that enzymes in the family of cytochrome P450 monooxygenases are involved in the conversion of these precursors to the final epoxide products. This study investigated the tissue specificity and sequence of these conversions by performing crude enzyme assays with protein extracts from male tissues. Furthermore, candidate P450 genes were selected by RNA-sequencing and co-expression analysis and the corresponding recombinant proteins tested for enzyme activity. To engineer the pheromone biosynthetic enzymes in plants, transient expression of the TPSs of both stink bugs was performed in Nicotiana benthamiana leaves. Both sesquipiperitol and (Z)-α-bisabolene were found to be produced and emitted from inoculated N. benthamiana leaves. Future work will implement stable transformation to engineer murgantiol biosynthesis in crucifer trap crops and develop similar approaches for pheromone engineering of other agricultural pests.
Master of Science
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Lyczakowski, Jan Jakub. "Biosynthesis and function of glucuronic acid substitution patterns on softwood xylan." Thesis, University of Cambridge, 2019. https://www.repository.cam.ac.uk/handle/1810/287949.

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Wood from coniferous trees is an important source of renewable biomass. It can contribute to provision of carbon neutral energy, biomaterials and housing for a growing population. Softwood is mainly composed of cellulose, galactoglucomannan, xylan and lignin. This thesis focuses on the biosynthesis and function of Glucuronic acid (GlcA) decorations on softwood xylan. Results demonstrate that this GUX (GlucUronic acid substitution of Xylan)-dependent xylan branching is critical for the maintenance of biomass recalcitrance in a model vascular plant Arabidopsis thaliana. Experiments employing in vitro and in planta activity assays show that conifer transcriptomes encode at least two distinct GUX enzymes which are active glucuronosyltransferases. Interestingly, these enzymes have different specific activities, with one adding evenly spaced GlcA branches and the other one being able to add consecutive decorations. It is possible that these different patterns of xylan branching may have an impact on ability of xylan to interact with cellulose fibrils. To investigate the role for xylan binding to cellulose, Arabidopsis mutant plants in which this interaction is lost were evaluated alongside transgenic mutant lines in which the interaction may be restored. Results of this analysis indicate that the presence of cellulose-bound xylan might have an influence on plant vasculature integrity and thus it may have an effect on plant growth and biomass properties. Moreover, further results indicate that some xylan cellulose interaction is likely to occur in cell wall macrofibrils which can be detected in softwood. Taken together, this thesis provides insights into the process of conifer xylan glucuronidation and the possible role these branches may be playing in the maintenance of softwood recalcitrance and mechanical properties. In addition to identifying potential mutagenesis targets for improving softwood processing, this work is a proof of concept for the use of GUX enzymes for in vivo and in vitro biosynthesis of novel xylan structures with potential industrial application.
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Rojas, Natalia Palacios. "Secondary metabolism in plants : molecular screening of floral diversity and studies of biosynthetic pathways in Lonicera tatarica and Catharanthus roseus." Thesis, University of East Anglia, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.323391.

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Broomhead, A. J. "Chemical and biochemical studies of tumour inhibitory aryl tetralin lignans." Thesis, University of Nottingham, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.235376.

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Dunn, Steven Mark. "The 5'-methylthioadenosine nucleosidase of pea (Pisum sativum)." Thesis, University of Exeter, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.314442.

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Häkkinen, Suvi T. "A functional genomics approach to the study of alkaloid biosynthesis and metabolism in Nicotiana tabacum and Hyoscyamus muticus cell cultures /." [Espoo, Finland] : VTT, 2008. http://www.vtt.fi/inf/pdf/publications/2008/P696.pdf.

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Decani, Yol Betul. "Extension Of Flower Longevity In Transgenic Plants Via Antisense Blockage Of Ethylene Biosynthesis." Master's thesis, METU, 2004. http://etd.lib.metu.edu.tr/upload/3/12605135/index.pdf.

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Ethylene (C2H4) is a very simple molecule, a gas, and has numerous effects on the growth, development and storage life of many fruits, vegetables and ornamental crops. In higher plants, ethylene is produced from L-methionine in essentially all tissues and ACC Synthase and ACC Oxidase are the two key enzymes in the biosynthesis of ethylene. The objective of the present study was to transform tobacco (Nicotiana tabacum L. cv. Samsun) plant with partial sequence of torenia acc oxidase gene in antisense and sense orientations via Agrobacterium-mediated gene transfer system, and to analyze its effect on ethylene production in transgenic plants. Six antisense and seven sense T0 putative transgenic lines were obtained and were further analyzed with several assays. Leaf disc assay and chlorophenol red assay under selection (75 mg/L kanamycin) revealed positive results compared to the non-transformed plant. T1 generations were obtained from all putative transgenic lines. PCR analysis and Northern Blot Hybridization results confirmed the transgenic nature of T1 progeny. Furthermore, ethylene amount produced by flowers were measured with gas chromatography, which resulted in an average of 77% reduction in S7 line and 72% reduction in A1 line compared with the control flowers. These results indicated that, transgenic tobacco plants carrying torenia acc oxidase transgene both in antisense and sense orientations showed reduced ethylene production thus a possibility of flower life extension.
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Sandager, Line. "Genes and enzymes involved in the biosynthesis of triacylglycerol in plants and yeast /." Alnarp : Swedish Univ. of Agricultural Sciences (Sveriges lantbruksuniv.), 2001. http://epsilon.slu.se/avh/2001/91-576-5832-3.pdf.

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38

Styer, Jean Christine. "Regulating Inositol Biosynthesis in Plants: Myo-Inositol Phosphate Synthase and Myo-Inositol Monophosphatase." Thesis, Virginia Tech, 2000. http://hdl.handle.net/10919/9870.

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Inositol is important for normal growth and development in plants. The regulation of the inositol biosynthetic enzymes, myo-inositol phosphate synthase (MIPS) and myo-inositol monophosphatase (IMP) was investigated. The specific aims of this research were (1) to develop a tool to study MIPS protein accumulation in a model plant system, Arabidopsis thaliana (At) and potentially other plant species and (2) to determine the spatial expression patterns of Lycopersicon esculentum IMP-2 (LeIMP-2) at the cellular level.

Myo-inositol phosphate synthase (mips) genes have been identified in plants, animals, fungi and bacteria. Alignment of the predicted amino acid sequences of AtMIPS-1, -2 and Glycine max MIPS (GmMIPS) indicated that AtMIPS-1 and GmMIPS are 87% identical, and AtMIPS-2 and GmMIPS are 89% identical. Based on these data, a Gmmips cDNA was fused at the N-terminus to a 6X histidine tag (5' GAC GAC GAC GAC GAC GAC 3'), cloned into an overexpression vector and overexpressed in E. coli. The fusion protein, HISMIPS, was extracted using denaturing conditions and purified using Ni2+ affinity chromatography. Anti-GmMIPS antiserum from rabbit detected the recombinant HISMIPS protein (76 kD), and GmMIPS (64 kD). Affinity purification by subtractive chromatography yielded anti-GmMIPS antibody that detected MIPS (66 kD) and a protein (34 kD) of unknown function. AtMIPS accumulated to high levels in unopened flowers, opened flowers, and immature siliques (6 mm in length or less), but was not detectable in bolts, cauline or rosette leaves.

The tomato inositol monophosphatase (Leimp) genes are a developmentally regulated multigene family. From analysis of sequences, Leimp-2 is intron-less and has the putative start site of translation located at +108 bp downstream from the putative start site of transcription. Investigation of the 5â UTR revealed the 3' end of a partial open reading frame (338 bp) highly homologous to the gene for calmodulin. Three light responsive elements and a cold responsive element were also identified in the 5' UTR.

Transgenic Leimp-2::uidA plants were produced using the existing construct of the Leimp-2 promoter fused to the uidA gene (J. Keddie, University of California at Berkeley). Seedlings were perserved and sectioned. Using histological techniques, the analysis of the Leimp-2 promoter::uidA transgenic seedlings revealed that the Leimp-2 promoter causes expression at the base of the shoot apex and within leaflets of the first set of fully expanded leaves. Further, Leimp-2 promoter expression was localized to epidermal and cortex cells on the abaxial side of the 1st and 2nd fully expanded compound leaves.

These studies of MIPS and IMP expression lay a foundation for a better understanding of the regulation of inositol biosynthesis in Arabidopsis, tomato, and other plant species.


Master of Science
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39

Abdul-Rahman, M. M. "Studies on the biosynthesis of ligands and their production in plant cell cultures." Thesis, University of Nottingham, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.381094.

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40

Vijh, Renu. "Role and compartmentation of ATP: citrate lyase in terpenoid and lipid biosynthesis in plants." Thesis, University of Hull, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.363184.

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41

Simon, Josiah William. "Molecular and structural studies on proteins involved in lipid biosynthesis in plants and bacteria." Thesis, Durham University, 2002. http://etheses.dur.ac.uk/4036/.

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Fatty acid synthesis in plants and most bacteria is catalysed by a type II, dissociable fatty acid synthetase, critically involving the participation of acyl-carrier protein (ACP) during synthesis and elongation reactions. This provides the substrate for glycerolipid biosynthesis, the first reaction of which in plants, is catalysed by a soluble glycerol-3-phosphate acyltransferase (G3PAT).A candidate gene for malonyl CoA:ACP transacylase (MCAT) from plants has been identified using homology to known MCAT sequences. A full-length cDNA was isolated from a Brassica napus embryo library and used to complement an E.coli mutant defective in MCAT activity, so providing proof of function. E.coli ACP has been investigated in attempts to obtain crystals which can be used in structural investigations to solve its structure via X-ray crystallography. Wild type butryl-ACP can be successfully crystallised and defracts to 2.0 Ǻ resolution, and is thus a candidate for solving the structure of the protein. The complete structure of butryl-ACP was elucidated following the introduction of new methionine residues into the protein using site directed mutagenesis, and the production of recombinant proteins containing selenomethionine as heavy metal derivatives. Squash recombinant G3PAT was over-expressed and used to obtain crystals which defract to1.8 Ǻ. In order to solve the phasing problem selected cysteine residues were removed from the protein, resulting in the identification of histidine 279 as an additional heavy metal binding site. The structure of the enzyme was solved to 1.8 Ǻ resolution and potential substrate binding sites have been modelled into it on the basis of the conserved H(X)(_4)D acyltransferase domain. The site for G3P binding has been confirmed following site directed mutagenesis and novel substrate selectivity has been introduced into the protein by PCR mediated mutagenesis. These studies have resulted in the first X-ray structure of two components of lipid biosynthesis, increased our understanding of the reactions they catalyse and successfully identified an authenticated cDNA for MCAT from plants.
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Thelander, Mattias. "Studies of molecular mechanisms integrating carbon metabolism and growth in plants /." Uppsala : Dept. of Plant Biology and Forest Genetics, Swedish Univ. of Agricultural Sciences, 2003. http://epsilon.slu.se/a432.pdf.

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43

Israelsson, Maria. "Gibberellin homeostasis and biosynthesis in relation to shoot growth in hybrid aspen /." Umeå : Dept. of Forest Genetics and Plant Physiology, Swedish Univ. of Agricultural Sciences, 2004. http://epsilon.slu.se/s307.pdf.

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Elbagory, Abdulrahman Mohammed Mohammed Nagy. "Green synthesis and characterization of gold nanoparticles from South African plants and their biological evaluations." University of the Western Cape, 2019. http://hdl.handle.net/11394/7011.

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Philosophiae Doctor - PhD
The field of nanotechnology continues to offer solutions for biotechnologists whose target is to improve the quality of life by finding new therapies to combat diseases. Gold nanoparticles (AuNPs) have been showing great potentials in many biomedical applications. The antibacterial activity of the AuNPs presents a therapeutic option for conditions caused by bacterial infections such as chronic wounds. Also, these versatile particles can offer solutions in the treatments of infectious diseases and can also be exploited as “smart” vehicles to carry drugs, such as antibiotics, for improved efficiency. Moreover, the anti-inflammatory activity of AuNPs makes them useful in the management of prolonged inflammation caused by bacterial infections. The synthesis of AuNPs can be achieved by variety of physical and chemical methods that have been successfully applied in labs and industry. Nonetheless, the drawbacks of these “conventional” methods in terms of high cost, adverse health side effects and incompatibility with the ecosystem cannot be overlooked. Thus, new safer and more cost-effective protocols have been reported for the synthesis of AuNPs. Plants have provided alternate synthesis methods in which the reducing capabilities of the phytochemicals, found in the aqueous plant extracts, can be used to chemically synthesize AuNPs from gold precursors. The biosynthesis and characterization of AuNPs from the phytochemicals of several South African plants is investigated in this study. The study also reports the optimization of the AuNPs biosynthesis by varying reaction conditions such as temperature and plant extracts’ concentrations. Furthermore, the study highlights the wound healing activity of the AuNPs synthesized from selected plants by investigating their antibacterial activity on bacterial strains known to cause chronic wounds. The ability of these AuNPs to carry ampicillin in order to enhance the antibacterial activity is also described herein. The cytotoxicity of the biosynthesized AuNPs was evaluated on human normal fibroblasts cells (KMST-6). Additionally, the immunomodulatory effect of the biosynthesized AuNPs on the cytokines production from macrophages and Natural Killer (NK) cells was examined. The study was successful to produce biocompatible and safe AuNPs synthesized from the tested aqueous plant extracts. The resulted AuNPs showed different physicochemical properties by varying the reaction conditions. The AuNPs exhibited antibacterial activity against several Gram-positive and Gram-negative bacteria. Also, ampicillin was successfully loaded on the biosynthesized AuNPs, which led to the formation of more antibacterial active conjugated AuNPs compared to the free AuNPs. The green synthesized AuNPs were also found to have anti-inflammatory responses as shown by the reduction of pro-inflammatory cytokines from immune cells. In vitro assays showed that the biogenic AuNPs were not toxic to KMST-6 cells. Overall, the data suggest that plant extracts produce biologically safe AuNPs with antibacterial and anti-inflammatory activities that can be exploited in the treatment of chronic wounds and in the management of chronic inflammation.
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45

Ge, Lingxiao. "The Export of Polyamines in Plants Is Mediated By a Novel Clade of Bidirectional Transporters." Bowling Green State University / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1430486987.

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46

Heinig, Uwe Herbert [Verfasser]. "Studies on the evolution of complex natural products biosynthetic pathways on basis of Taxol biosynthesis in plants and endophytic fungi / Uwe Herbert Heinig." Aachen : Hochschulbibliothek der Rheinisch-Westfälischen Technischen Hochschule Aachen, 2012. http://d-nb.info/1024800075/34.

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47

Dowdle, John. "Aspects of the control of L-ascorbic acid biosynthesis via the Smirnoff-Wheeler pathway in plants." Thesis, University of Exeter, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.410807.

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48

Groh, Chloé. "Deciphering the interactions between plants and bacteria in the context of isoprenoid biosynthesis in Arabidopsis thaliana." Thesis, Strasbourg, 2019. http://www.theses.fr/2019STRAJ079.

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Les isoprénoïdes constituent une grande classe de molécules que l'on retrouve chez tous les organismes vivants et qui sont impliquées dans divers processus biologiques. Le but de ma thèse était de déterminer si ces molécules pouvaient être impliquées dans les interactions entre plantes et bactéries. J'ai travaillé sur des plantes Arabidopsis thaliana sauvages, et sur deux mutants affectés dans chacune des deux voies de biosynthèse des isoprénoïdes existant chez les plantes supérieures. Un inventaire des communautés constituant le microbiote de ces plantes a mis en évidence certaines bactéries dont l’abondance varie dépendamment du génotype, malgré l'existence d'un microbiote commun entre plantes sauvages et mutantes. De plus, il a été démontré que les plantes affectées dans la voie de biosynthèse plastidiale, dite du méthylérythritol phosphate (MEP), sont plus sensibles que les plantes sauvages à l’infection par Pseudomonas syringae, un phytopathogène bien connu. Ensemble, ces résultats suggèrent que les isoprénoïdes peuvent jouer un rôle dans les interactions entre plantes et bactéries. D’autre part, 230 souches interagissant avec A. thaliana ont été isolées au laboratoire, parmi lesquelles certaines ont été testées pour déterminer leur effet sur le phénotype et la résistance des plantes à P. syringae, ainsi que l’effet des isoprénoïdes sur ces souches
Isoprenoids are a large class of molecules found in all living organisms that are implicated in diverse biological processes. The aim of my thesis was to decipher if they could be involved in the interactions between plants and bacteria. Therefore, I worked on Arabidopsis thaliana wild-type and mutants, altered in the two isoprenoid biosynthesis pathways occurring in higher plants. An inventory of the communities interacting with wild-type and mutants highlighted some differentially abundant bacteria, despite the existence of a core microbiota. In addition, plants affected in the plastidial biosynthesis pathway, referred as the methylerythritol phosphate (MEP) pathway, have been shown to be more affected than wild-type by Pseudomonas syringae, a well-known phytopathogen. Together, these results suggest that isoprenoids may play a role in the interactions between plants and bacteria. In addition, 230 strains were isolated from A. thaliana in the laboratory, among which some of them were tested for their impact on the plant fitness and resistance to P. syringae, and for the impact on isoprenoids on these strains
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49

Zhong, Yujuan, and 钟玉娟. "Functional analysis of green algal {221}-carotene ketolases and metabolic engineering of astaxanthin biosynthesis in higher plants." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B47145730.

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Astaxanthin (3,3'-dihydroxy-β-carotene-4,4'-dione) is a ketocarotenoid that is beneficial for human health due to its ability of boosting immune function and preventing tumor formation. The biosynthesis of astaxanthin is, however, limited only to a few organisms. The burgeoning demand for natural astaxanthin has attracted much recent interest in extending the carotenoid pathway of higher plants to astaxanthin by expressing a microbial β-carotene ketolase (BKT). One major challenge of engineering an astaxanthin pathway in plants is the low astaxanthin content achieved. Five green microalgae including Chlamydomonas reinhardtii, Chlorococcum sp., Neochloris wimmeri, Protosiphon botryoides and Scotiellopsis oocystiformis were selected with enhanced function for astaxanthin biosynthesis. The products of the BKT cDNAs from the algae are similar in sequence to the BKT from Haematococcus pluvialis (ca 70% amino acid identity). Based on an Escherichia coli system, the BKT enzymes were shown to exhibit various efficacies in converting zeaxanthin into astaxanthin with Chlamydomonas BKT exhibiting the highest conversion rate (ca 85%). To investigate if the function-enhanced Chlamdomonas BKT (CRBKT) has advantages over other algal BKTs in triggering astaxanthin biosynthesis in higher plants, the CrBKT, together with the BKTs from Chlorella zofingiensis (CzBKT) and H. pluvialis (HpBKT3) was expressed in Arabidopsis thaliana. Transgenic Arabidopsis expressing the CrBKT developed orange leaves which accumulated astaxanthin up to 2 mg g-1 dry weight. In contrast, the expression of CzBKT resulted in much lower content of astaxanthin (0.24 mg g-1 dry weight), whereas HpBKT3 was unable to mediate synthesis of astaxanthin in Arabidopsis. Similarly, overexpression of CrBKT in tobacco also resulted in the massive accumulation of astaxanthin in leaves (1.60 mg g-1 DW). Taken all together, it can be concluded that ketolating zeaxanthin efficiently is essential for high production of astaxanthin in transgenic plants. Tomato is an important food crop with high amounts of carotenoids in its fruit. To investigate if tomato fruit can serve as a bio-factory for astaxanthin production, the CrBKT was overexpressed in three genotypes of tomato. All transgenic tomato plants developed brown red leaves that accumulated canthaxanthin rather than astaxanthin as a major carotenoid, resulting from the poor catalytic activity of the endogenous BHY1 toward canthaxanthin. To overcome this problem, CrBKT and HpBHY, the best pair of genes catalyzing the formation of astaxanthin in β- carotene-producing E. coli, were coexpressed in tomato. Canthaxanthin was efficiently converted to astaxanthin, resulting in a massive accumulation of astaxanthin in leaf (3.12 mg g-1) and fruit (16.1 mg g-1) with enhanced total carotenoid capacities of 1.7-fold in leaf and 16.6-fold in fruit. Moreover, the over-production of astaxanthin in fruit enhanced its antioxidant capacity 3-5-fold and vitamin C 2-fold, although it did not affect growth and development. In summary, the Chlamydomonas BKT is proven to be superior to other sources of BKT/CrtW enzymes in triggering astaxanthin biosynthesis in plants. By coexpressing a pair of well-cooperating BKT and BHY genes, the transgenic B-type tomato could accumulate commercially attractive amounts of the high-value astaxanthin in its fruit. This study highlights the potential of higher plants to be engineered as cell factories for producing the high-value astaxanthin.
published_or_final_version
Biological Sciences
Doctoral
Doctor of Philosophy
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50

Stander, Emily Amor. "Unraveling the transcriptome of Aspalathus linearis (Rooibos) towards identification of novel genes involved in polyphenol biosynthesis." University of the Western Cape, 2019. http://hdl.handle.net/11394/6921.

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Philosophiae Doctor - PhD
South Africa (SA) is home to one of the six floral kingdoms of the world, and hosts a very diverse flora comprising an astonishing ~30,000 species. Herbal medicines play an important role in many of the diverse cultures of this country. Yet, agricultural production systems for most of these species are missing, and medicinal plants are usually collected in the wild. The endemic medicinal plants of SA produce a wide range of rare medicinally active compounds, which could be developed into drugs. Knowledge on the genes involved in the biosynthesis of these compounds could not only promote establishment of plant production systems, but also their biotechnological exploitation. Transcriptomics has been revolutionized by Next Generation Sequencing technologies, which can cost-efficiently provide a lot of information on plant genes and biosynthetic pathways. This thesis focuses on the establishment of methodologies for high-throughput plant transcriptome research, including: 1) harvesting plant material suitable for high-quality RNA analysis from distant locations, 2) high-throughput, and inexpensive biochemical sample screening, 3) extraction of high-quality RNA from recalcitrant, polysaccharide- and polyphenol rich plant material, and 4) biocomputational analysis of Illumina sequencing data, including quality control and pre-processing of data, de novo assembly of reads, protein prediction and functional transcriptome annotation. Rooibos (Aspalathus linearis) was chosen as the pilot plant, because it is one of the few indigenous SA medicinal plants that has been successfully cultivated as a commercial crop. It produces a wide range of phenolic compounds with health promoting properties (e.g. aspalathin and a phenylpropenoic acid glucoside with scientifically verified antidiabetic and cardioprotective effects). In the course of this study, seven rooibos transcriptomes were produced, assembled and functionally annotated, providing a first extensive dataset for identification of genes associated with economically important traits such as medicinal compound production, rooibos growth type characteristics and stress resistance.
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