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Post-harvest seedcoat darkening is a major problem in many pulses, including common bean (</i>Phaseolus vulgaris</i> L.). In some bean market classes, such as pinto, beans that have a darkened seedcoat are discounted in the market place as it is assumed that the beans are old and will be hard-to-cook (HTC). Pinto genotypes that darken more slowly than conventional pinto beans would be more desirable and have been identified in the bean breeding program at the University of Saskatchewan. <p>To study the slow-darkening trait, a quick, reliable, and inexpensive screening method that would not affect seed germination would be beneficial. Three potential protocols to accelerate seedcoat darkening were examined. The greenhouse protocol was conducted in the greenhouse by placing the bean seeds in polybags with a 1 cm2 piece of moistened felt. For the UV light protocol, bean seeds were placed 10 cm below an UV lamp which had a wavelength of 254 nm. For the cabinet protocol, bean seeds were placed in a cabinet set at 30¢ªC, 80% relative humidity, and full fluorescent lights. Color measurements were taken routinely using a Hunter Lab colorimeter. All three methods were successful in distinguishing darkening beans from slow-darkening beans although the UV light protocol was considered to be superior to the greenhouse and cabinet protocol as the UV light protocol was quick, consistent over years, and the most economical. Unlike the greenhouse and the cabinet protocols, the UV light protocol did not affect seed germination following accelerated darkening. <p>The stability of the slow-darkening trait was further investigated in genotype by environment (g x e) studies across different indoor and outdoor environments. In the g x e study across different field environments, it was found that prior to accelerated seedcoat darkening the g x e interaction was significant. Following accelerated seedcoat darkening, environment and genotype were both significant and g x e was not. The slow-darkening genotypes had lighter seedcoats than the darkening genotypes and those field sites that had more favorable weather had lighter seedcoats. For the g x e study across indoor and outdoor environments, when the genotypes were split into either slow-darkening or darkening, the g x e interaction was not significant and the slow-darkening genotypes had lighter seedcoats. <p>Genetic control of the slow darkening trait was determined. For crosses between slow-darkening genotypes and CDC Pintium, the F2 populations segregated 3 darkening : 1 slow-darkening with distinct bimodal distribution. This indicated that seedcoat darkening was controlled by a single gene and darkening was dominant over slow-darkening. For both slow-darkening by slow-darkening crosses, the F2 populations¡¯ L* values were unimodal, normal distributions, indicating there may be modifying genes for the slow-darkening trait.

Post-harvest seedcoat darkening is a major problem in many pulses, including common bean (</i>Phaseolus vulgaris</i> L.). In some bean market classes, such as pinto, beans that have a darkened seedcoat are discounted in the market place as it is assumed that the beans are old and will be hard-to-cook (HTC). Pinto genotypes that darken more slowly than conventional pinto beans would be more desirable and have been identified in the bean breeding program at the University of Saskatchewan. <p>To study the slow-darkening trait, a quick, reliable, and inexpensive screening method that would not affect seed germination would be beneficial. Three potential protocols to accelerate seedcoat darkening were examined. The greenhouse protocol was conducted in the greenhouse by placing the bean seeds in polybags with a 1 cm2 piece of moistened felt. For the UV light protocol, bean seeds were placed 10 cm below an UV lamp which had a wavelength of 254 nm. For the cabinet protocol, bean seeds were placed in a cabinet set at 30¢ªC, 80% relative humidity, and full fluorescent lights. Color measurements were taken routinely using a Hunter Lab colorimeter. All three methods were successful in distinguishing darkening beans from slow-darkening beans although the UV light protocol was considered to be superior to the greenhouse and cabinet protocol as the UV light protocol was quick, consistent over years, and the most economical. Unlike the greenhouse and the cabinet protocols, the UV light protocol did not affect seed germination following accelerated darkening. <p>The stability of the slow-darkening trait was further investigated in genotype by environment (g x e) studies across different indoor and outdoor environments. In the g x e study across different field environments, it was found that prior to accelerated seedcoat darkening the g x e interaction was significant. Following accelerated seedcoat darkening, environment and genotype were both significant and g x e was not. The slow-darkening genotypes had lighter seedcoats than the darkening genotypes and those field sites that had more favorable weather had lighter seedcoats. For the g x e study across indoor and outdoor environments, when the genotypes were split into either slow-darkening or darkening, the g x e interaction was not significant and the slow-darkening genotypes had lighter seedcoats. <p>Genetic control of the slow darkening trait was determined. For crosses between slow-darkening genotypes and CDC Pintium, the F2 populations segregated 3 darkening : 1 slow-darkening with distinct bimodal distribution. This indicated that seedcoat darkening was controlled by a single gene and darkening was dominant over slow-darkening. For both slow-darkening by slow-darkening crosses, the F2 populations¡¯ L* values were unimodal, normal distributions, indicating there may be modifying genes for the slow-darkening trait.

The ascomycete fungus Magnaporthe oryzae, causative agent of rice blast disease, poses a threat to global food security, destroying enough rice to feed 60 million people each year. Characterization of the host-pathogen interaction between rice and M. oryzae is critical, as better understanding of the system may lead to better disease control strategies. The sequenced genome and repertoire of molecular tools available have made M. oryzae an ideal model system for understanding general plant-pathogen interactions as well. The objective of this dissertation was to characterize the M. oryzae homologs of Histoplasma capsulatum RYP (Required for Yeast Phase) genes that are required for transition to the parasitic phase. H. capsulatum is a human pathogen that undergoes a dimorphic switch from filamentous to yeast cell growth at 37°C, the host body temperature. Four H. capsulatum RYP genes were identified in a forward genetic screen to identify genes required for entry into the yeast phase. RYP1 is a member of the Gti1_Pac2 family, which contains previously characterized regulators of dimorphic switching. RYP2 and RYP3 are homologs of vosA and velB, members of the Velvet family, best characterized in Aspergillus nidulans, where they coordinate morphological differentiation with secondary metabolism. RYP4 is a zinc binuclear cluster protein, a main class in the zinc finger transcription factor family. Deletion of the M. oryzae RYP1 homolog, RIG1 (Required for Infectious Growth), resulted in a non-pathogenic mutant on susceptible rice cultivars, even upon removal of the host penetration barrier. Δrig1 was blocked in the transition to infectious hyphal growth, similar to H. capsulatum ryp1, which could not transition to the yeast phase. Deletion mutants of M. oryzae RYP2, RYP3, and RYP4 homologs were similar to the wild type in somatic growth and pathogenicity indicating that although RIG1 is a pathogenicity factor conserved in plant and animal pathogens, such conservation does not apply to all of the RYP pathogenicity genes identified in H. capsulatum. Δrig1 is the first M. oryzae mutant known to be blocked in production of primary infection hyphae. Overall, the study suggests limited parallels exist in phase transition of fungal pathogens of plants and animals.

Corn (Zea mays L.) seedlings possessed a high degree of tolerance to growth inhibitory effects of pre-emergent application of 2-chloro-N-isopropylacetanilide (propachlor), squash (Cucurbita maxima Duchesne) was intermediate in its response to the herbicide followed by oats (Avena sativa L.), lambsquarters (Chenopodium album L.) and redroot (Amaranthus retroflexus L.). Root growth was more severely retarded by lower concentrations of propachlor than was stem development. Propachlor inhibited cell division in onion (Allium cepa L.) root tips. Aberrations in the mitotic behavior, such as contraction of chromosomes, was also induced. Cell elongation, in particular auxin induced cell elongation of oat coleoptile , was inhibited in direct proportion to propachlor concentration. Concomitant with the inhibition of squash seedling growth and development, propachlor prevented the normal senescence of squash cotyledons. Cell wall expansion and breakdown and the utilization of proteinaceous and lipid reserves in the cotyledons were inhibited by propachlor treatment. Amino acid activation, both in vivo as well as in vitro, was significantly reduced in squash whereas in corn, amino acid activation was inhibited only in vitro. This inhibition of amino acid activation is postulated to be the primary mechanism of propachlor's herbictdal action. This inhibition would prevent the formation of proteinaceous compounds including (1) the enzyme complexes which would break down storage material , and (2) alter cell walls to allow cell elongation.

The buildup of salt in soils is a major abiotic stress that affects agricultural productivity, limiting the growth and yield of most crop species which cannot tolerate even modest levels of salinity (glycophytes). Genetic variability for salt tolerance exists as some plants (halophytes) have adapted to environments with high levels of salt. Understanding how salt tolerance has been acquired in halophytic species will be an important part of strategies to improve the ability of crops to grow in saline soils. The CALCINEURIN B-LIKE10 (AtCBL10) calcium sensor was identified as a component of salt signaling in the glycophyte Arabidopsis thaliana (A. thaliana) based on hypersensitivity of the Atcbl10 mutant to salt. When A. thaliana is grown in the presence of salt, AtCBL10 interacts with the AtSOS2 protein kinase to activate the AtSOS1 sodium/proton exchanger, resulting in the removal of sodium ions from the cytosol. Eutrema salsugineum (E. salsugineum), a halophytic relative of A. thaliana, has two CBL10 genes (EsCBL10a and EsCBL10b). In this research, the duplication of CBL10 in E. salsugineum was characterized and the functions of EsCBL10a and EsCBL10b in salt tolerance were determined. My analyses indicate that the coding sequences of EsCBL10a and EsCBL10b are highly conserved, as they share 85% nucleotide identity. An analysis of transcript structure indicates transcripts from EsCBL10a and EsCBL10b loci are alternatively spliced, but in distinct ways. My results suggest that EsCBL10a and AtCBL10 likely share the ancestral genomic position, while EsCBL10b might have moved to a different genomic region, and that the duplication took place prior to the divergence of expanded Lineage II species. The expression patterns of EsCBL10a and EsCBL10b are different; EsCBL10b transcript is high in shoots and low in roots while EsCBL10a transcript is detectable in both tissues. Preliminary analysis of E. salsugineum lines with reduced expression of EsCBL10a and EsCBL10b suggest that both genes might play a role during growth in the presence of salt, but that these roles are distinct.

Strawberry production in the United States is almost entirely done in open-fields. Recently, interest in off-season strawberry production using controlled environment (CE) systems such as greenhouses and soilless cultivation has increased in the US. However, strawberry production in greenhouses is relatively new in North America and available information about greenhouse strawberry production is limited. Plant physiological responses to the environment must be well understood to maximize production using CE systems. In the present research, photoperiodic and photosynthetic responses of strawberry plants in greenhouse were studied. To evaluate photoperiodic response eight cultivars of strawberry widely cultivated in North America were subjected to varied photoperiods under an average daily temperature of 17 °C. Short-day (SD) cultivars included 'Radiance', 'F-127', 'Shuksan' and 'Chandler', and day-neutral/ever-bearing (DN/EB) cultivars included 'Albion', 'Portola', 'Monterey' and 'San Andreas'. SD cultivars were subjected to treatments of 11-h, 12-h, 13-h and 14-h photoperiod for 8 weeks. DN/EB cultivars were subjected to separated treatments of 8-h, 11-h, 14-h and 17-h photoperiod for up to 10 weeks. After 8 and 10 weeks of photoperiodic treatments in SD and DN/EB cultivars respectively, shoot apical meristems (SAM) were observed under microscope and classified into one of twelve developmental stages (Indices: 0-11). All SD cultivars examined showed a critical photoperiod between 13 h and 14 h. DN/EB cultivars 'San Andreas', 'Albion' and 'Monterey' presented facultative long-day response with positive correlation between SAM developmental indices and photoperiod after 8 weeks of treatment. 'Portola' showed non-significant influences of photoperiodic treatments in flower primordial development, suggesting day neutral response. However, further experiments must be conducted to confirm cultivar responses and identify possible interactions between photoperiod and temperature. The effect of light intensity and plant sink/source balance on strawberry plant photosynthesis was also investigated. Measurements of leaf net photosynthetic rate (Pn), stomatal conductance, intercellular CO2 concentration (Ci) and transpiration rate under near-saturated photosynthetically photon flux of 1,000ï ­mol m-2 s-1 and ambient CO2 concentration in strawberry cultivars 'Albion' and 'Nyoho'; and tomato cultivar 'Speedella'(comparison purposes) were conducted monthly in greenhouse during May 2014, and monthly from September 2015 through May 2016 (winter/spring production season). Hourly measurements were recorded hourly from 9 AM to 4 PM. Potential source strength was determined by the number of leaves and the daily light integral (DLI, 400-700 nm) and sink load from flowers and fruits was represented as the number of flowers and fruits. Seasonal changes in daily maximum Pn were observed, as well as diurnal change in Pn in both strawberry cultivars. A significant positive correlation was determined between the estimated ratios of sink/source and the slopes representing diurnal linear decline of Pn. Also, Pn was negatively correlated with Ci but not significantly correlated with vapor pressure deficit (VPD) in greenhouse, suggesting diurnal decline in Pn was likely due to negative feedback of photosynthesis caused by unbalance of sink and source, and not to water stress from high VPD. The photosynthetic capacity as affected by seasonal changes in greenhouse environment and its diurnal change as affected by sink/source balance could help develop more effective practices in CE strawberry production to maximize production. Also, photoperiodic response revealed in this study for American strawberry cultivars will specify conditions to induce flowering in these economically important cultivars off-season.

Nicotiana benthamiana is now an established platform for molecular farming, the production of biopharmaceuticals in plants. Infiltration with Agrobacterium tumefaciens (agroinfiltration) is commonly used to transiently express one or multiple transgenes in N. benthamiana leaves. Agroinfiltrated N. benthamiana is a flexible and scalable recombinant protein (RP) production platform, but is impeded by low RP yields. Plant proteases can degrade RPs and thus limit RP accumulation. To inform, design and implement strategies for enhancing RP accumulation, I present four papers about proteases and protease inhibitors in agroinfiltrated N. benthamiana. First, I investigated the transcriptome, extracellular proteome and active secretome to understand the plant response to agroinfiltration and investigate the expressed proteases. I show that an extracellular immune response is mounted at the expense of photosynthesis. Comprehensive annotation and monitoring uncover a large, diverse repertoire of proteases in agroinfiltrated leaves, indicating that broad-range depletion of protease activity may be required to enhance RP accumulation. Second, I reviewed the literature on multifunctional plant protease inhibitors (PIs) and grouped them into three types of multifunctional PIs that evolved independently. Third, I screened candidate PIs and discovered that three new, unrelated PIs enhance RP accumulation. I present universal elements of the RP degradation machinery, uncovering new questions on our understanding of the protease network that degrades RPs. Fourth, I identified targets of SlCYS8, a PI that enhances RP accumulation. The target proteases of SlCYS8 are implicated in RP degradation and the high specificity of SlCYS8 can be used to study their role in other processes. By elucidating the immune response to agroinfiltration, by uncovering the N. benthamiana protease repertoire and by providing new tools to deplete the activity of specific proteases, this thesis makes a relevant contribution to both basic plant research and molecular farming.

Kabuli chickpea (<i>Cicer arietinum</i> L.) has two leaf types, the fern and unifoliate. Yield potential is limited for kabuli chickpea in Saskatchewan. It is limited by a short-season, a semi-arid environment, and end-of-season rainfall. Manipulating plant population, and choosing chickpea cultivars with the best leaf type for biomass production, radiation interception and yield for the early, middle, or late growth season, may increase chickpea yield. Therefore, the objectives of this study were: to (i) determine the relationship between leaf type and key growth parameters of six chickpea cultivars varying in leaf morphology at moderate and high plant population densities; (іі) to characterize the reaction of the fern and unifoliate leaf to altered canopy light environments. Different light environments were created by 50% defoliation at vegetative growth, first flower, and 50% shading from vegetative growth to first flower, as well as two light enrichment treatments initiated at the first flower and pod formation stages.<p> Fern leaf cultivars exhibited higher maximum light interception, seasonal cumulative intercepted radiation and a higher harvest index compared to unifoliate leaf cultivars. However, both leaf type canopies had less than 95% light interception for most of the season. The fern and unifoliate leaf type contributed to similar radiation use efficiency in three out of four location-years. In addition, fern leaf cultivars produced significantly higher seed yield than cultivars with unifoliate leaves.<p>Plant density influenced growth parameters. For example, the 45 plants m-2 treatment had a higher harvest index than the 85 plants m-2 treatment, in two location-years, while both population treatments were similar in the other two location-years. Yield of chickpea was increased by higher plant population in only one location-year, but was not significantly affected by plant population in the other location-years. The effect of canopy light environment manipulation on chickpea yield depended on the stages of plant development when they were applied. Defoliation at vegetative growth and first flower had no effect on yield. However, plants responded significantly to the 50% shade treatment; the crop growth rate, harvest index and yield were less in the shaded treatment compared to the control. Shading also increased plant height. Light enrichment treatments increased the yield. However, the degree of yield increase was greater when light enrichment occurred at first flower, than at the later stage of pod formation. These results highlighted the importance of the amount of irradiance during the flowering stage. It was concluded that chickpea breeders should select lines with fern leaves for improved radiation interception when breeding cultivars for semiarid short-season environments such as in Saskatchewan. Management and breeding practices should ensure that the crop can make efficient use of the solar radiation at flowering to maximize yield. Improvement at the canopy and subsequent yield level is yet to be made in Saskatchewan environments by increased light interception, increased growth before flowering, and increased and stable harvest index.

Meadow bromegrass (<i>Bromus riparius</i> Rehm.) is an important forage grass in western Canada. Economical seed production is critical to its use. Seed yield usually declines rapidly after two to three seed crops. Field and growth chamber experiments were conducted to determine the influence of (a) residue removal and N fertilization on tiller density and size, panicle density, silvertop incidence, seed yield and stand longevity, (b) filler size and stand age on panicle production, and (c) daylength and temperature during primary and secondary induction on panicle production. Removing residue after harvest and applying N (100 kg ha-1 ) increased yield from 200 to 450 kg ha-1 compared to not removing residue or adding N in the second seed crop. The difference between treated and untreated plots was only 30 to 90 kg ha-1 in the third seed crop. The increase was related to an increase in panicle production, however, the correlation between panicle number and seed yield was low. Silvertop incidence (% of panicles affected) increased as the stand aged, but removing residue after harvest reduced silvertop. In pot studies, the percentage of plants that produced panicles increased as tiller basal diameter increased from one mm to three mm, regardless of the age of the stand. However, fewer large tillers were observed in older stands. Large tillers from a four-yr-old stand produced fewer particles than large tillers from a two-yr-old stand indicating that tiller size alone is not responsible for the decline in panicle production. Panicle production increased as the temperature during primary induction decreased. However, daylength during primary induction had no effect on panicle production. Varying temperature or daylength during secondary induction had no effect on panicle production; panicles were produced in 85% of plants regardless of temperature, and 67 to 77% of the plants regardless of daylength. In conclusion, residue removal after harvest and N fertilization improve seed yield in young meadow bromegrass stands. However, these practices were not effective in prolonging seed yield beyond two to three seed crops. Drought, winter injury, competition amongst tillers and silvertop incidence all play a role in reducing seed production.