| 1. |
- Zubair, Muhammad
(författare)
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Genetic variation, biochemical contents and wound healing activity of Plantago major
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Plantago major L. (greater plantain, common plantain) has been used as a wound healing remedy in different parts of the world for centuries. Different bioactive compounds have been proposed to contribute to the wound healing properties of this plant. The present study was undertaken to investigate the impact of some genetic and environmental factors on the wound healing activity of common plantain. Seeds of P. major were collected from five populations in different parts of Sweden, and were germinated and grown in a greenhouse. As expected for an inbreeding species, RAPD analyses demonstrated considerable between-population variation but very sparse within-population and within-subpopulation variation. Six major phenolic compounds were encountered in samples of P. major, four of which were identified for the first time in this thesis; PLMA 1–PLMA 4. Between-population and sub-population differences in the contents of these chemical compounds showed no correlation with RAPD-based estimates of genetic relatedness. The contents of these compounds differed greatly between different plant organs of P. major. The highest concentration of plantamajoside and PLMA 2 was found in leaves whereas the highest concentration of verbascoside was found in flower stalks and seeds. Contents were significantly higher in freeze-dried leaf samples compared to samples dried at higher temperatures. Both water and ethanol-based extracts of P. major leaves stimulated the cell proliferation and migration in an in vitro scratch assay, and also showed anti-inflammatory activity in an in vitro NF-kB assay with oral epithelial cell cultures. Similarly, these extracts stimulated wound healing activities in ex vivo tests using detached pig ears. Further breeding efforts aimed at developing P. major as a crop plant, and medicinal research aimed at elucidating and optimizing extracts with wound healing properties, are thus warranted.
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| 2. |
- Wang, Yang
(författare)
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Discovery and investigation of glycoside hydrolase family 5 enzymes with potential use in biomass conversion
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Glycoside hydrolases (GHs) cleave glycosidic bonds in glycoconjugates, oligosaccharides and polysaccharides such as cellulose and various hemicelluloses. Mannan is a major group of hemicelluloses. In higher plants, they usually serve as storage carbohydrates in seeds and tubers or as structural polysaccharides cross-linking with cellulose/lignin in cell walls. In industrial fields, this renewable biomass component can be used in various areas such as production of biofuels and health-benefit manno-oligosaccharides; and mannan degrading enzymes, especially mannanases, are important molecular tools for controlling mannan polysaccharides properties in biomass conversion. In this thesis, the evolution, substrate specificity and subfamily classification of the most important GH family, i.e., glycoside hydrolase family 5 (GH5), are presented providing a powerful tool for exploring GH5 enzymes in search for enzymes with interesting properties for sustainable biomass conversion. Additionally, three GH5_7 mannanases from Arabidopsis thaliana (AtMan5-1, AtMan5-2 and AtMan5-6) were investigated in the present study. Bioinformatics tools, heterologous expression, and enzymology were applied in order to reveal the catalytic properties of the target enzymes, increase understanding of plant mannanase evolution, and evaluate their potential use in biomass conversion. This approach revealed: (1) AtMan5-1 exhibits mannan hydrolase/transglycosylase activity (MHT), (2) AtMan5-2 preferably degrades mannans with a glucomannan backbone, and (3) AtMan5-6 is a relatively thermotolerant enzyme showing high catalytic efficiency for conversion of glucomannan and galactomannan making this plant mannanase an interesting candidate for biotechnological applications of digesting various mannans. Moreover, these studies suggest an evolutionary diversification of plant mannanase enzymatic function.
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| 3. |
- Roos, Jonas
(författare)
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Verticillium longisporum and plant immunity responses in Arabidopsis
- 2014
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Verticillium spp. are soil-borne ascomycete fungi belonging to a subgroup of Sordariomycetes, and the three major plant pathogens Verticillium longisporum, V. dahliae and V. albo-atrum cause disease on numerous plant species worldwide. In Sweden, V. longisporum poses a threat to Brassica oilseed crops, and is thus emphasized in this thesis. Here the early immune responses to V. longisporum in the model plant Arabidopsis and recent data on the V. longisporum genome are presented. Three genes of importance in the Arabidopsis–V. longisporum interaction were studied. The genes were identified via transcriptome and single nucleotide polymorphism (SNP) analysis. RabGAP22, a RabGTPase-regulating protein, was found to contribute to V. longisporum resistance. Pull-down assays revealed SERINE:GLYOXYLATE AMINOTRANSFERASE (AGT1) as an interacting partner during V. longisporum infection and the two proteins were shown to co-localize in the peroxisomes. Unexpectedly, a role for RabGAP22 was also found in stomatal immunity. The monoterpene synthase TPS23/27 was on the other hand found to contribute to fungal invasion, by triggering germination of V. longisporum conidia. The third gene codes for a nitrate/peptide transporter, NPF5.12. Pull-down experiments and fluorescent imaging revealed interaction between NPF5.12 and a major latex protein family member, NPFBP1. Implications in plant immunity processes of these three genes are further discussed. The genomes of two Swedish V. longisporum isolates were sequenced and found to have a size of approximately 70 Mb and harbor ~21,000 protein-coding genes. Initial analyses revealed that 86% of the V. longisporum genomes are shared with V. dahliae and V. albo-atrum, with a high extent of gene duplications. Large numbers of proteins were predicted to contain secretion motifs, and this group of proteins is presumed to play major roles in the interactions with V. longisporum host plants. In conclusion, this thesis work has revealed new fungal and plant host genes and thereby laid the basis for new plant breeding and disease protection strategies.
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| 4. |
- Berglund, Jennie
(författare)
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Wood Hemicelluloses - Fundamental Insights on Biological and Technical Properties
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Hemicelluloses are a group of heterogeneous polysaccharides representing around 30 % of wood where the dominating types are xylans, glucomannans and xyloglucans. Hemicelluloses complex molecular structure makes it difficult to understand the relationship between structure and properties entirely, and their biological role is not yet fully verified. Additionally, hemicelluloses are sensitive to chemical processing and are not utilized to their full potentials for production of value-added products such as materials, additives to food and pharmaceutical products, etc. Increased knowledge regarding their functions is important for the development of both processes and products. The aim with this work has therefore been to increase the fundamental understanding about how the structure and properties of wood hemicelluloses are correlated, and properties such as flexibility, interaction with cellulose, solubility, resistance to chemical-, thermal-, and enzymatic degradation have been explored.Molecular dynamics (MD) simulations were used to, in detail, study the structures found in wood hemicelluloses. The flexibility was evaluated by comparing the impact of backbone sugars on the conformational space and also the impact of side groups was considered. Based on the conformational space of backbone glycosidic linkages the flexibility order of hemicelluloses in an aqueous environment was determined to be: xylan > glucomannan > xyloglucan. Additionally, the impact of xylan structure on cellulose interaction was evaluated by MD methods.Hemicelluloses were extracted from birch and spruce, and were used to fabricate different composite hydrogels with bacterial cellulose. These materials were studied with regards to mechanical properties, and it was shown that galactoglucomannans mainly contributed to an increased modulus in compression, whereas the most significant effect from xylan was increased strain under uniaxial tensile testing. Besides, other polysaccharides of similar structure as galactoglucomannans were modified and used as pure, well defined, models. Acetyl groups are naturally occurring decorations of wood hemicelluloses and can also be chemically introduced. Here, mannans with different degrees of acetylation were prepared and the influence of structure on solubility in water and the organic solvent DMSO were evaluated. Furthermore, the structure and water solubility influenced the interaction with cellulose. Acetylation also showed to increase the thermal and biological stability of mannans.With chemical pulping processes in mind, the degradability of spruce galactoglucomannans in alkaline solution were studied with regards to the structure, and the content of more or less stable structural regions were proposed.
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| 5. |
- Calleja-Rodriguez, Ainhoa
(författare)
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Quantitative Genetics and Genomic Selection of Scots pine
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The final objective of tree improvement programs is to increase the frequency of favourable alleles in a population, for the traits of interest within the breeding programs. To achieve this, it is crucial to decompose the phenotypic variance accurately into its genetic and environmental components in order to obtain a precise estimation of genetic parameters and to increase genetic gains. The overall aim of this thesis was to increase the accuracy of genetic parameter estimation by incorporating new quantitative genetics models to the analysis of multiple traits in multiple trials of Scots pine, and to develop a genomic selection protocol to accelerate genetic gain. Factor analysis was incorporated to multivariate multi-environment analyses and it allowed to evaluate up to 19 traits simultaneously. As a result, precise patterns of genotype-by-environment interactions (G E) were observed for tree vitality and height; moreover, it was possible to detect the main driver of the G E: differences in temperature sum among sites. Traditional quantitative trait loci (QTL) analysis of phenotypic data was compared with the detection of QTL with estimated breeding values (EBV) for the first time in a three generation pedigree and, as outcome, it was noticed that if a QTL was associated to a EBV and to a phenotypic trait, the proportion of variance explained by the QTLEBV was higher than the QTL-phenotype. Additionally, several QTL were detected across several ages, which may make them suitable as candidates for early selection. Genomic selection (GS) could aid to reduce the breeding cycle by shortening the periods of progeny field testing, and consequently increasing genetic gains per year. Genomic predictions, including additive and non-additive effects through different prediction models were compared with traditional pedigree-based models; it was seen an overestimation of genetic parameters for pedigree-based models, even larger when nonadditive effects could not be discerned from additive and residual effects. Prediction accuracies and abilities of the genomic models were sufficient to achieve higher selection efficiencies and responses per year varying between 50-90% by shortening 50% the breeding cycle. For the selection of the top 50 individuals, higher gains were estimated if non-additive effects are incorporated to the models (7 – 117%).
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| 6. |
- Leonova, Svetlana
(författare)
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Lipids in seeds of oat (Avena spp.), a potential oil crop : content, quality, metabolism, and possibilities for improvement
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- New oil crops with high yield and oil content are urgently needed. Oat is the only cereal that accumulates substantial amount of lipids in the endosperm. This gives it potential as an oil crop, which could address both nutritional and environmental concerns of modern society. To develop oat with increased oil content, thorough investigations on its lipid metabolism and tools for genetic manipulations are needed. Analyses of lipids in seeds of wild and cultivated oat species revealed variation in wild oat accessions in both oil content and quality. This variation should allow for development of new oat varieties for diverse applications. However, to develop oat as an oil crop, a level of 20% of the seed dry weight as oil is required and the range of oil content in the studied accessions was 4-10%. Another feature which showed almost no deviation was the amount of omega 3 fatty acid, -linolenic acid (0.6-2.1%). To develop oat varieties with ultra-high seed oil and increased content of omega 3 fatty acids, a biotechnological approach is preferred to conventional breeding methods. Lipids in oat endosperm have been reported to exist, not enclosed in oil bodies, but as non-structured oil smear. In my study, I also observed oil smears, a phenomenon probably correlated with the reduced amount of oil body associated proteins in the endosperm, as revealed by microscopic and staining methods. This was supported experimentally by SDS-PAGE separation of oil-body proteins and immunoblotting and immunolocalization with antibodies against a 16 kD oil body protein. Biochemical studies on lipid mobilization during oat seed germination demonstrated efficient utilization of oil reserves from the starchy endosperm; these results were corroborated by microscopy. An oat cultivar which is capable of accumulating high amounts of oil in the endosperm was shown to be also efficient in utilizing these reserves upon germination. Agrobacterium-mediated transformation is one of the methods for modification of oat oil content and/or its quality. The results of the analyses aimed at development of an Agrobacterium-mediated oat transformation protocol are presented in this thesis.
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| 7. |
- Nemesio Gorriz, Miguel
(författare)
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Molecular responses against Heterobasidion annosum s.l. in Picea abies
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Norway spruce [Picea abies (L.) Karst.] is a main tree species in European forests and is important both ecologically and economically. The root rot fungus Heterobasidion annosum sensu lato (s.l.) is the main P. abies pathogen. Including resistance in breeding programs would help mitigating the impact of the pathogen but knowledge regarding defense mechanisms in P. abies needs a better understanding. The work within this thesis intended to expand the existing knowledge on P. abies resistance mechanisms focusing on hormone signaling, flavonoid biosynthesis and its transcriptional regulation. I found that jasmonic acid is the major hormone controlling defense signaling pathways in P. abies. Furthermore, we validated a candidate gene, PaLAR3, as a resistance marker for H. annosum s.l. in P. abies. PaLAR3 encodes an enzyme responsible for the synthesis of (+) catechin, which showed a fungistatic effect on H. parviporum. Analysis of genetic diversity revealed two allelic lineages of PaLAR3 that showed significant differences in fungal resistance and (+) catechin content that were explained by dissimilarities in inducibility. We studied the role of PaNAC03, a transcription factor that is associated with H. annosum s.l. infection. PaNAC03 not only showed repression of multiple genes including PaLAR3, but bound only to the promoter of one of the PaLAR3 allelic lineages explaining at least partly the differences in allelic expression that were observed. Finally, we identified a full repertoire of members of a MYB/bHLH/WDR transcription factor complex in Norway spruce, which showed differences in protein interactions and expression patterns, and also in ability to control the expression of genes in the flavonoid biosynthetic pathway including PaLAR3.
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| 8. |
- Zhu, Tianqing
(författare)
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Regulation of embryo development in Norway spruce by WOX transcription factors
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In seed plants, the apical-basal axis of the plant body is established during early embryogenesis. Major regulatory genes of the apical-basal axis formation belong to the WUSCHEL-RELATED HOMEOBOX (WOX) gene family of transcription factors. The spatiotemporal expression pattern and the molecular role of the WOX genes has mainly been studied in the angiosperm model plant Arabidopsis (Arabidopsis thaliana). Similar information in conifers is limited. The aim of my thesis has been to characterize WOX genes in Norway spruce (Picea abies) and to elucidate the function of WOX genes expressed during embryo development. We cloned 11 WOX homologs from Norway spruce and examined their phylogenetic relationship to WOX genes from other species. The phylogenetic analyses showed that the major diversification within the WOX gene family took place before the gymnosperm-angiosperm split. PaWOX8/9, PaWOX2 and PaWOX3, which are expressed in embryos, were selected for further studies. PaWOX8/9 and PaWOX2 are highly expressed in early and late embryos, and PaWOX3 is highly expressed in mature embryos. Functional studies were performed in RNAi lines where the genes were down-regulated. Embryos in PaWOX8/9 RNAi lines showed a disturbed apical-basal patterning caused by abnormal orientation of the cell division plane at the basal part of the embryonal mass. In PaWOX2 RNAi lines, vacuolated cells differentiated on the surface of the embryonal mass and the embryos failed to form a proper protoderm. Down-regulation of PaWOX3 disturbed lateral margin outgrowth in cotyledons and needles. Taken together, our results indicate that WOX8/9, WOX2 and WOX3 exert evolutionarily conserved functions during embryo development. We can therefore conclude that the regulatory networks of embryo development are at least partly conserved between angiosperms and gymnosperms.
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| 9. |
- Ahmadi Afzadi, Masoud
(författare)
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Genetic variation in resistance to fungal storage diseases in apple : inoculation-based screening, transcriptomics and biochemistry
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Apple is one of the economically and culturally most important fruit crops and has many health-related benefits. Apple production is, however, sensitive to several fungal diseases including blue mold, caused by Penicillium expansum. Problems are more pronounced in organic production or in countries where postharvest application of fungicides is prohibited. To limit or overcome this problem, many studies have been focused on investigations of the mechanism of resistance/tolerance. No major gene(s) have as yet been identified, but quantitatively inherited traits, some of which are related to fruit texture and content of chemical compounds, have been shown to affect the ability of cultivars to withstand storage diseases. In the present thesis, inter-cultivar variation in terms of resistance to fungal storage diseases was investigated at two locations, i.e. Balsgård in Sweden and Njøs in Norway. The association of harvest date, fruit firmness and softening with lesion decay was investigated on large sets of cultivars. The contribution of four fruit texture-related genes (Md-ACO1, Md-ACS1, Md-Exp7 and Md-PG1) in explaining the fruit texture characteristics was examined. Fruit content of chemical compounds with a potential impact on disease resistance was also investigated, and finally the regulation of apple genes upon fungal infection was studied in order to identify candidate genes responsible for disease resistance. Inoculation-based screening indicated large variation across the investigated cultivars in terms of blue mold and bitter rot susceptibility. Harvest date and softening rate of fruits during storage had a large impact on resistance to fungal diseases, thus cultivars with moderate to firm fruits that soften comparatively little during storage could withstand the fungal infection comparatively well. Softening rate is, in its turn, closely associated with harvest date whereas four fruit texture-related genes had lower predictive power than expected. Quantifying the chemical compounds in the fruit samples revealed that some of these compounds, especially flavonols and procyanidin B2, could contribute to resistance against blue mold, whereas contents of malic acids or total titratable acidity had considerably less impact. Differential expression of FLS, LDOX, and CHS genes involved in biosynthesis of flavonoids and PGIP, TT10, WAK1 and CTL1 genes related to cell wall structure indicate the importance of fruit characteristics and biochemical compounds in the resistance mechanism.
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| 10. |
- Pham, Toan Duc
(författare)
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Analyses of genetic diversity and desirable traits in sesame (Sesamum indicum L., Pedaliaceae) : implication for breeding and conservation
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Sesame (Sesamum indicum L., Pedaliaceae) is a traditional oil crop cultivated in Vietnam and Cambodia. It is known as the king of oil seeds in Vietnam due to the high oil content (50-60%) in its seed. Yet, the insufficient genetic information regarding Vietnamese and Cambodian sesame populations is limiting the access to useful traits present among adapted landraces in this region. The purpose of this study was to characterize various sesame accessions to gain information that could help design strategies for future breeding program and conservation of this crop in the two countries. Morphological and molecular markers as well as oil content and quality analyses were employed to evaluate sesame accessions from different sources. High genetic variation was found among populations of sesame collected in Vietnam and Cambodia. The two type of markers, morphological and molecular, were both useful in analyzing the extent of genetic diversity in sesame and the result of these analyses will help to better understand the genetic diversity and relationship within and among populations. Overall, the sesame accessions included in the study showed a correlation with their geographical origins such that accessions from the same region tended to have higher genetic similarity as compared to those from different regions. However, when cluster analysis was applied to evaluate the genetic relationship, some sesame accessions were found not to be grouped based on geographical origins. This contrasting result could perhaps be a result from the exchange over time, of sesame germplasm, between farmers across the regions. The results from morphological and oil content analyses showed that several sesame accessions in Southern Vietnam and Cambodia displayed a good potential for high seed yield and oil content. Overall, the studies in this thesis provide important insights into the populations of sesame in Vietnam and Cambodia and constitute a set of useful background information that can be used as a basis for future breeding strategy and improvement of sesame in this region.
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