by Li Wing Yen Francisca. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 94-105). / Abstracts in English and Chinese. / Thesis committee --- p.i / Statement --- p.ii / Abstract --- p.iii / Chinese Abstract --- p.v / Acknowledgemnets --- p.vii / Abbreviations --- p.ix / Table of contents --- p.xii / List of figures --- p.xvi / List of tables --- p.xvii / Chapter 1. --- General Introduction / Chapter 1.1 --- Introduction to oxidative stress / Chapter 1.1.1 --- Introduction to Reactive Oxygen Species --- p.1 / Chapter 1.1.2 --- Major sites of ROS production / Chapter 1.1.2.1 --- Chloroplast --- p.4 / Chapter 1.1.2.2 --- Mitochondria --- p.4 / Chapter 1.2 --- Regulation of intercellular ROS content in plant cells / Chapter 1.2.1 --- Enzymatic defense ofROS --- p.6 / Chapter 1.2.1.1 --- Superoxide dismutases --- p.6 / Chapter 1.2.1.2 --- "Ascorbate peroxidase, Glutathione reductase and the Ascorbate-Glutathione cycle" --- p.7 / Chapter 1.2.1.3 --- Catalase --- p.11 / Chapter 1.2.1.4 --- Alternative oxidase --- p.11 / Chapter 1.2.2 --- Non-enzymatic / Chapter 1.2.2.1 --- Ascorbate and Glutathione --- p.12 / Chapter 1.2.2.2 --- α-tocopherol --- p.12 / Chapter 1.3 --- "Salt, dehydration and oxidative stress" / Chapter 1.3.1 --- Oxidative stress is induced when plants were under salt stress --- p.13 / Chapter 1.3.2 --- Oxidative stress is induced when plants were under dehydration stress --- p.14 / Chapter 1.4 --- ROS scavenging: the road to achieve multiple-stress tolerance? --- p.16 / Chapter 1.5 --- Purple acid phosphatase and its relationship with oxidative stress in plants / Chapter 1.5.1 --- General introduction to plants purple acid phosphatase (PAP) --- p.20 / Chapter 1.5.2 --- Purple acid phosphatases that found to be involved in ROS scavenging in plants --- p.21 / Chapter 1.6 --- Previous studies in GmPAP3 --- p.23 / Chapter 1.7 --- Hypothesis and significance of this project --- p.25 / Chapter 2. --- Materials and methods / Chapter 2.1 --- Materials / Chapter 2.1.1 --- "Plants, bacterial strains and vectors." --- p.26 / Chapter 2.1.2 --- Chemicals and reagents --- p.27 / Chapter 2.1.3 --- Commercial kits --- p.28 / Chapter 2.1.4 --- Primers and adaptors --- p.29 / Chapter 2.1.5 --- Equipments and facilities used --- p.31 / Chapter 2.1.6 --- "Buffer, solution, gel and medium" --- p.31 / Chapter 2.1.7 --- Software --- p.31 / Chapter 2.2 --- Methods / Chapter 2.2.1 --- Molecular techniques / Chapter 2.2.1.1 --- Bacterial cultures for recombinant DNA and plant transformation --- p.32 / Chapter 2.2.1.2 --- Recombinant DNA techniques --- p.32 / Chapter 2.2.1.3 --- "Preparation and transformation of DH5α, DE3 and Agrobacterium competent cells" --- p.33 / Chapter 2.2.1.4 --- Gel electrophoresis --- p.36 / Chapter 2.2.1.5 --- DNA and RNA extraction --- p.37 / Chapter 2.2.1.6 --- Generation of single-stranded DIG-labeled PCR probes --- p.38 / Chapter 2.2.1.7 --- Testing the concentration of DIG-labeled probes --- p.40 / Chapter 2.2.1.8 --- Northern blot analysis --- p.40 / Chapter 2.2.1.9 --- PCR techniques --- p.41 / Chapter 2.2.1.10 --- Sequencing --- p.42 / Chapter 2.2.2 --- Plant cell culture and transformation / Chapter 2.2.2.1 --- Arabidopsis thaliana --- p.43 / Chapter 2.2.2.2 --- Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) cells --- p.44 / Chapter 2.2.3 --- Growth and treatment conditions for plants / Chapter 2.2.3.1 --- Growth and salt treatment condition of soybean samples for gene expression studies of GmPAPS --- p.45 / Chapter 2.2.3.2 --- Root assay of GmPAP3 transgenic Arabidopsis thaliana --- p.46 / Chapter 2.2.4 --- "Immunolabeling, mitochondria integrity, ROS detection and confocal microscopy" / Chapter 2.2.4.1 --- Immunolabeling of GmPAP3-T7 transgenic cell lines --- p.47 / Chapter 2.2.4.2 --- Mitochondria integrity --- p.48 / Chapter 2.2.4.3 --- Detection of Reactive oxygen species (ROS) --- p.48 / Chapter 2.2.4.4 --- Confocal microscopy --- p.49 / Chapter 2.2.4.5 --- Images processing and analysis --- p.49 / Chapter 2.2.5 --- Statistical analysis --- p.50 / Chapter 3. --- Results / Chapter 3.1 --- "Expression of GmPAP3 was induced by NaCl stress, oxidative stress, and dehydration stress" --- p.51 / Chapter 3.2 --- Establishment of GmPAP3-T7 fusion transgenic cell lines / Chapter 3.2.1 --- Subcloning of GmPAP3-T7 into the binary vector system W104 --- p.53 / Chapter 3.2.2 --- Transformation of W104-GmPAP3-T7 into tobacco BY-2 cells --- p.56 / Chapter 3.3 --- Establishment of GmPAP3 trangenic cell lines / Chapter 3.3.1 --- Subcloning of GmPAP3 into the binary vector system W104 --- p.58 / Chapter 3.3.2 --- Transformation of W104-GmPAP3 into tobacco BY-2 cells --- p.58 / Chapter 3.4 --- Establishment of GmPAP3 transgenic Arabidopsis thaliana / Chapter 3.4.1 --- Transformation of W104-GmPAP3 into Arabidopsis thaliana --- p.61 / Chapter 3.5 --- Colocalization of GmPAP3 with MitoTracker-orange --- p.66 / Chapter 3.6 --- Effect of expressing GmPAP 3 on mitochondria integrity of BY-2 cells under NaCl and dehydration stress. --- p.71 / Chapter 3.7 --- Effect of expressing GmPAP3 on ROS production in BY-2 cells under salt and PEG treatment --- p.75 / Chapter 3.8 --- Effect of expressing GmPAP3 in Arabidopsis thaliana under salt stress --- p.81 / Chapter 4. --- Discussion / Chapter 4.1 --- Gene expression profile of GmPAP3 --- p.83 / Chapter 4.2 --- Subcellular localization of GmPAP3 --- p.84 / Chapter 4.3 --- Functional tests of GmPAP 3 transgenic BY-2 cells / Chapter 4.3.1 --- GmPAP3 could protect the plant cells' mitochondria integrity when under salt and dehydration stress --- p.86 / Chapter 4.3.2 --- Expressing GmPAPS in tobacco BY-2 cells were able to reduce the production ofROS under salt and dehydration stresses --- p.88 / Chapter 4.4 --- Functional tests of GmPAP3 transgenic Arabidopsis --- p.91 / Chapter 5. --- Conclusion and perspectives --- p.92 / References --- p.94 / Appendix I: Restriction and modifying enzymes --- p.106 / Appendix II: Chemicals --- p.107 / Appendix III: Commercial kits --- p.111 / Appendix IV: Equipments and facilities used --- p.112 / "Appendix V: Buffer, solution, gel and medium formulation" --- p.113

One of the most important photosynthetic enzymes in a plant is ribulose-1,5¬bisphosphate carboxylase/oxygenase (Rubisco), which plays a key role in carbon fixation. Degradation of this enzyme leads to decreased carbon fixation and poor photosynthetic performance by the plant. It is therefore of interest to investigate possible ways of protecting this enzyme during stress conditions in order to generate plants that would perform better under extreme climates. In this study the effect of an expressed, exogenous rice cysteine proteinase inhibitor (OCI) in transformed tobacco plants on Rubisco stability/content under chilling and senescence was investigated. Results showed that there is no significant protective role for exogenous OCI on the degradation/content of Rubisco when tobacco plants were exposed to chilling. This result was found using native gel-based quantification procedures, as well as immuno-blotting, spot densitometric analysis, and a radioactive quantification assay as analysis techniques. The study, however, provided evidence for protection of Rubisco against degradation by expression of OCI under a more severe stress condition, such as senescence using native gel-based quantification procedures as detection techniques. Tobacco plants were also transformed with a newly designed vector allowing expressed OCI to be transported to the chloroplast. Failure to detect so far any OCI-¬expressing transformed plants and the idea that delay of senescence could prove beneficial to farmers by providing a more nutrient-dense crop with higher tolerance against stress-induced cell death are discussed. / Dissertation (MSc(Botany))--University of Pretoria, 2006. / Plant Science / unrestricted

Plant productivity is greatly affected by environmental stresses such as drought, salinity and insect herbivory. Plants respond and adapt to these stresses by exhibiting physiological as well as biochemical changes at the cellular and molecular levels in order to survive. Expression of a variety of genes which encode numerous membrane transporters have been demonstrated to be induced by these stresses in a variety of plants. The nutritional status of plants is controlled by these transporters, which are regulated by the transcription of the corresponding genes. In spite of these adverse stress effects on agricultural yield, only a few studies have focused on gene transcriptional and translational regulation of membrane transporters during environmental stress situations. Rice, like other plants, contains a number of sucrose transporters encoded by a family of genes. However, detailed knowledge of their roles, localization and regulation during environmental stress conditions is lacking. Bioinformatic tools were used to identify putative cis-acting regulatory elements that may be involved in the regulation of rice and Arabidopsis thaliana sucrose transporters. The possible cis-acting regulatory elements were predicted by scanning genomic sequences 1.5 kbp upstream of the sucrose transporter genes translational start sites, using Plant CARE, PLACE and Genomatix Matinspector professional data bases. Several cis-acting regulatory elements that are associated with plant development, plant hormonal regulation and stress response were identified, and were present in varying frequencies within the 1.5 kbp of 5′ regulatory region. The putative cis-acting regulatory elements that possibly are involved in the expression and regulation of sucrose transporter gene families in rice and Arabidopsis thaliana during cellular development or environmental stress conditions were identified as: A-box, RY, CAT, Pyrimidine-box, Sucrose-box, ABRE, ARF, ERE, GARE, Me-JA, ARE, DRE, GA-motif, GATA, GT-1, MYC, MYB, W-box, and I-box. Expression analysis was used to elucidate the role of rice (Oryza sativa L. cv Nipponbare) sucrose transporter (OsSUT) genes during drought and salinity treatments of three week old rice plants ( at four leaf stage) over a 10 days. Among the five rice OsSUT genes identified, only OsSUT2 was observed to be progressively up-regulated during drought and salinity treatments, while OsSUT1, OsSUT4 and OsSUT5 were expressed at low levels, and OsSUT3 showed no detectable transcript expression. Sucrose transport will be essential to meet the cellular energy demands and also for osmoprotectant activities during drought and salinity stresses. It therefore indicates that OsSUT2 which facilitates transport of sucrose from photosynthetic cells will be III essential for rice plants to cope with drought and salinity stresses, and cultivars with a higher OsSUT2 expression should be able to tolerate these environmental stresses better. The role of OsSUT in assimilate transport during rusty plum aphids (Hysteroneura setariae; Thomas) infestation on the leaves of three week old rice (Orzya sativa L. cv Nipponbare) cultivar plants, over a time-course of 1 to 10 days of treatments, was also examined by combination of gene expression and β-glucuronidase (GUS) reporter gene analysis. Real Time PCR analysis of the five OsSUT genes revealed that the expression of OsSUT1 was progressively up-regulated during the course of aphid infestation. OsSUT2 and OsSUT4 expression were comparatively low in both the control and treated plants. OsSUT5 showed no clear difference in transcript expression in both control and treated plants, while no detectable transcript expression of OsSUT3 could be found. The up-regulation of OsSUT1 gene was verified at protein level by western blot analysis in both the control and treated plants. OsSUT1 protein expression was found to increase with time during aphid infestation. A similar trend was noticeable in the control plants, however at a lower expression level. These demonstrate that the cellular expression of OsSUT1is regulated by both developmental and environmental factors. OsSUT1-promoter:::GUS reporter gene expression was observed within the vascular parenchyma and/or companion cells associated with phloem sieve elements of the large and small bundles in the phloem tissues of the flag leaf blade regions where feeding aphids were confined, which progressively increased with time of infestation. It is suggested that OsSUT1 may primarily play an essential role in phloem transport of assimilate to wounded tissues from adjacent health tissues or may be involved in the retrieval of assimilate back into the phloem to minimize loss caused by the infestation. Some OsSUT1-promoter:::GUS expression was also found in the metaxylem at 10 days after infestation, which could signify a recovery system in which sucrose lost into the xylem as a result of aphids feeding are retrieved back into the phloem through the vascular parenchyma. This was supported by the exposure of cut ends of matured OsSUT1-promoter:::GUS rice plant leaf to 2% sucrose solution. OsSUT1-promoter:::GUS expression was observed within the protoxylem, xylem and phloem parenchyma tissues. This indicates that sucrose translocating within the xylem tissues are retrieved into the phloem via the OsSUT1 localized within the parenchyma tissues. In conclusion, the differential expression and regulation of rice (Orzya sativa L. cv Nipponbare) sucrose transporters as reported here suggest that OsSUT2 and OsSUT1 were constitutively expressed compared to other rice sucrose transporters during drought and salinity, and rusty plum aphids (Hysteroneura setariae; Thomas) infestation stresses respectively. Thus, the expression and regulation of the sucrose transporters could be related to the physiological and nutritional requirements of the cells during plant developmental or environmental stress state that allows their differential expression.

Water scarcity induced by drought, temperature, and salinity has plagued agricultural sustainability in recent years with unprecedented revenue losses, raising concerns for worldwide food security. Recent studies have revealed unique botanical response mechanisms to combat water related stress, namely the expression of proteins known as the dehydrins. Dehydrin proteins have been shown to serve various intracellular protective functions. The gene for a SK5 type dehydrin from the arctic plant Cerastium arcticum (CaDHN) was introduced into tobacco plants and water deficit tolerance was evaluated. Plants overexpressing CaDHN displayed improved tolerance to salt stress, but no improvement was observed under drought stress. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

Thesis submitted in fulfilment of the requirements for the degree Master of Technology: Horticulture in the Faculty of Applied Sciences at the Cape Peninsula University of Technology, 2012 / This article based thesis includes two individual studies evaluating the role of silicon (Si) in mitigating the negative effects that are associated with sodium chloride (NaCl) induced toxicity in lettuce (Lactuca sative L. 'Eish!').

Vegetation status is a key indicator of the ecosystem condition in a particular area. The study objective was about the estimation of leaf nitrogen (N) as an indicator of vegetation water stress using vegetation indices especially the red edge based ones, and how leaf N concentration is influenced by various environmental factors. Leaf nitrogen was estimated using univariate and multivariate regression techniques of stepwise multiple linear regression (SMLR) and random forest. The effects of environmental parameters on leaf nitrogen distribution were tested through univariate regression and analysis of variance (ANOVA). Vegetation indices were evaluated derived from the analytical spectral device (ASD) data, resampled to RapidEye. The multivariate models were also developed to predict leaf N. The best model was chosen based on the lowest root mean square error (RMSE) and higher coefficient of determination (R2) values. Univariate results showed that red edge based vegetation index called MERRIS Terrestrial Chlorophyll Index (MTCI) yielded higher leaf N estimation accuracy as compared to other vegetation indices. Simple ratio (SR) based on the bands red and near-infrared was found to be the best vegetation index for leaf N estimation with exclusion of red edge band for stepwise multiple linear regression (SMLR) method. Simple ratio (SR3) was the best vegetation index when red edge was included for stepwise linear regression (SMLR) method. Random forest prediction model achieved the highest leaf N estimation accuracy, the best vegetation index was Red Green Index (RGI1) based on all bands with red green index when including the red edge band. When red edge band was excluded the best vegetation index for random forest was Difference Vegetation Index (DVI1). The results for univariate and multivariate results indicated that the inclusion of the red edge band provides opportunity to accurately estimate leaf N. Analysis of variance results showed that vegetation and soil types have a significant effect on leaf N distribution with p-values<0.05. Red edge based indices provides opportunity to assess vegetation health using remote sensing techniques. / Environmental Sciences / M. Sc. (Environmental Management)

MSCAGR (Plant Production) / Department of Plant Production / Chickpea (Cicer arietinum L.) is an excellent utilizer of residual soil moisture in agricultural ecosystems. However, its seed quality and hence reproduction is constrained by water stress, seed size and storage conditions. This study was carried out at the University of KwaZulu- Natal (UKZN), Pietermaritzburg Campus. It was conducted to evaluate the performance of chickpea genotypes (Desi-K, Saina-K and ICCV-K) with different seed sizes on seedling emergence (i), seed ageing effect on seed quality and imbibition of genotypes produced under water stressed and non-stressed conditions (ii), and (iii) the effect of water stress during seed development on sugars and protein accumulation, germination and seed vigour. Pot experiments were conducted under glasshouse / tunnel conditions at the Controlled Environment Facilities (CEF). The experiment for objective 1 was laid out as a single factor in completely randomized design (CRD). Data on emergence rate, final hypocotyl and complete emergence was collected. The small seeded Desi-K showed higher and faster emergence compared to medium sized Saina-K and large seeded ICCV-K. In the experiment of the second objective, seeds of the three genotypes were first obtained by production under water stressed and non-stressed growing conditions. They were then aged for 0, 1, 3, 5, or 7 days at 41 ºC and 100% relative humidity to form a 2 x 3 x 5 (water levels x genotypes x ageing) factorial design. Data was collected on germination percentage (GP), mean germination time (MGT), electrical conductivity (EC), tetrazolium chloride test (TZ) and imbibition weight. Seed ageing caused progressive loss of seed viability and vigour in all genotypes, which resulted in lower GP, delayed MGT, reduced TZ staining, cell death and high solute leakage from the seeds produced under the two water regimes. However, the effect was more severe under water stressed conditions. In the experiment for objective 3, seeds of all three genotypes were larger when grown under non-stressed condition compared to those under water stressed condition. These larger seeds had higher seed viability and germination percentage but lower electrical conductivity and mean germination time. Stressed seeds had higher soluble sugars than non-stressed seeds. It was deduced that irrigation during seed development reduces the final sugars and protein content but increases the seed size and physiological quality parameters allied to production of chickpea. Therefore, water provision to chickpea crop is critical during seed development. / NRF