Master of Science / Department of Agronomy / Tesfaye Tesso / Cold temperature stress is an important abiotic constraint to grain sorghum production in temperate regions. In the United States, low temperature in late spring and early fall has limited sorghum production to a narrow growing period. Deployment of cold tolerance traits may widen this window and hence contribute to increased production. The objectives of this study were (1) to determine the effect of early and mid-season cold temperature stress on growth, phenology and yield components of sorghum, and identify key traits that are most sensitive to cold stress at seedling and flowering stages, and (2) to identify new sources of cold tolerance for use in breeding programs. Series of controlled environment (greenhouse/growth chamber) and field experiments were carried out. Three sorghum genotypes of variable response, Shan Qui Red (tolerant), SRN39 (susceptible) and Pioneer 84G62 (unknown) were subjected to cold (15/13ºC day/night) and normal (25/23ºC day/night) temperature at seedling (Experiment I) and flowering (Experiment II) stages. The genotypes were planted in a greenhouse using a 5L polytainer pots. Each pot consisted of a single plant and each plot was represented by three pots. A split-plot design with three replications was used in both experiments with temperature regimes as main plots and genotypes as sub-plots. Three days after emergence, experiment I plants were moved to the growth chamber and subjected to the designated temperature treatments. For experiment II, the treatments were assigned at heading stage immediately before anthesis had begun. The treatments lasted 10 d in both experiments. Data were collected on seedling characteristics and leaf chlorophyll content in experiment I, days to flowering, maturity, and yield components in both experiments, and anthesis duration in experiment II. For the field experiment, 150 sorghum germplasm collections of potential cold tolerance along with tolerant and susceptible checks were evaluated for emergence and seedling traits under early planting (April 13) at soil temperature of 20.1/13.4 ºC max/min. The normal temperature treatment was applied by planting at regular season (May 26) at soil temperature of 30.0/20.4ºC max/min. Twenty-four genotypes selected based on field emergence and seedling vigor were further screened under controlled environment. Early-season stress significantly reduced leaf chlorophyll content, all seedling traits (height, vigor and dry weight), and also delayed flowering and maturity. But it had no effect on final leaf number, plant height and yield components. Genotypic response to early stress was significant for all traits with the susceptible checks having the lowest score for all seedling traits. Mid-season cold stress prolonged anthesis duration, delayed maturity and highly reduced all yield components. Several genotypes among the 150 had higher seedling vigor and emergence than the tolerant check, Shan Qui Red. In conclusion, reduced seedling vigor as a result of early stress had no effect on final yield provided that stand establishment was not compromised while mid season stress is damaging to yield. The wide genetic variation for the traits indicates the potential for improvement of cold tolerance in sorghum.

Some Arabidopsis lesion mimic mutants (LMM) show alterations in abiotic stress responses as well as pathogen resistance. cpr22 is a LMM which has a mutation in cyclic nucleotide-gated ion channels, is a typical LMM exhibiting elevated levels of salicylic acid (SA), spontaneous cell death, constitutive expression of defense genes, and enhanced resistance to various pathogens in an SA dependant manner. cpr22 defense responses are suppressed in high humidity and enhanced by low humidity. To investigate environmental effects, microarray analyses were conducted. Expression of several genes related to abscisic acid (ABA) signaling was altered and ABA levels increased in cpr22 after humidity shift. Furthermore, significant alterations in ABA-related phenotypes were observed. Double mutant analysis with nahG plants indicated that alterations in ABA signaling were attributable to elevated SA levels. These results suggest a negative effect of SA on ABA signaling/abiotic stress responses during the activation of defense responses.

Using the short-lived radiotracer 13N, we examined the hypothesis that toxic, futile ammonia/ammonium (NH3/NH4+) fluxes at high external concentrations are mediated by ammonia-transporting aquaporins in roots of intact barley (Hordeum vulgare L.) plants. Effects of the aquaporin inhibitors zinc, copper, mercury, gold, silver, hydrogen peroxide, propionic acid, and nitrogen gas supported this hypothesis. Further tests with these inhibitors showed that changes in plant water potential and water content could be linked to NH3/NH4+ fluxes. An increase in external pH, causing an increase of NH3 in the nutrient solution, resulted in large increases of 13N influx, which can only be explained in energetic terms if the transported solute is neutrally charged. Taken together, the evidence here strongly supports the proposed hypothesis.

Using the short-lived radiotracer 13N, we examined the hypothesis that toxic, futile ammonia/ammonium (NH3/NH4+) fluxes at high external concentrations are mediated by ammonia-transporting aquaporins in roots of intact barley (Hordeum vulgare L.) plants. Effects of the aquaporin inhibitors zinc, copper, mercury, gold, silver, hydrogen peroxide, propionic acid, and nitrogen gas supported this hypothesis. Further tests with these inhibitors showed that changes in plant water potential and water content could be linked to NH3/NH4+ fluxes. An increase in external pH, causing an increase of NH3 in the nutrient solution, resulted in large increases of 13N influx, which can only be explained in energetic terms if the transported solute is neutrally charged. Taken together, the evidence here strongly supports the proposed hypothesis.

Some Arabidopsis lesion mimic mutants (LMM) show alterations in abiotic stress responses as well as pathogen resistance. cpr22 is a LMM which has a mutation in cyclic nucleotide-gated ion channels, is a typical LMM exhibiting elevated levels of salicylic acid (SA), spontaneous cell death, constitutive expression of defense genes, and enhanced resistance to various pathogens in an SA dependant manner. cpr22 defense responses are suppressed in high humidity and enhanced by low humidity. To investigate environmental effects, microarray analyses were conducted. Expression of several genes related to abscisic acid (ABA) signaling was altered and ABA levels increased in cpr22 after humidity shift. Furthermore, significant alterations in ABA-related phenotypes were observed. Double mutant analysis with nahG plants indicated that alterations in ABA signaling were attributable to elevated SA levels. These results suggest a negative effect of SA on ABA signaling/abiotic stress responses during the activation of defense responses.

Two projects were undertaken to study the non-energy conserving alternative pathway present in the plant mitochondrial ETC. In the first project, a tobacco AOX2 gene was cloned and characterized. AOX2 showed tissue specificity in expression and could not be induced by common stresses. In the second project I carried out a physiological characterization of transgenic tobacco plants with increased or decreased expression of AOX1 subjected to cold stress. Under non-stress condition, a strong inverse relationship between levels of AOX1 and levels of oxidative damage was observed, while after cold treatment AOX1 transgenic lines and WT showed more complicated and differential responses in aspects of oxidative damage and the capacity of antioxidant system. I also discovered that the pool sizes of monosaccharides after temperature shift were proportional to AOX1 levels. These results indicated that AOX1 might have crucial but complex impacts on ROS balance and carbon metabolism during cold stress.

Two projects were undertaken to study the non-energy conserving alternative pathway present in the plant mitochondrial ETC. In the first project, a tobacco AOX2 gene was cloned and characterized. AOX2 showed tissue specificity in expression and could not be induced by common stresses. In the second project I carried out a physiological characterization of transgenic tobacco plants with increased or decreased expression of AOX1 subjected to cold stress. Under non-stress condition, a strong inverse relationship between levels of AOX1 and levels of oxidative damage was observed, while after cold treatment AOX1 transgenic lines and WT showed more complicated and differential responses in aspects of oxidative damage and the capacity of antioxidant system. I also discovered that the pool sizes of monosaccharides after temperature shift were proportional to AOX1 levels. These results indicated that AOX1 might have crucial but complex impacts on ROS balance and carbon metabolism during cold stress.

Master of Science / Department of Biology / Jesse B. Nippert / The Konza Prairie in northern Kansas, USA contains over 550 vascular plant species; of which, few have been closely studied. These species are adapted to environmental stress as imposed by variable temperature, precipitation, fire, and grazing. Understanding which plant traits relate to drought responses will allow us to both predict drought tolerance and potential future shifts in plant community composition from changes in local climate. Morphological and physiological measurements were taken on 121 species of herbaceous tallgrass prairie plants grown from seed in a growth chamber. Gas exchange measurements including maximum photosynthetic rate, stomatal conductance to water vapor, and intercellular CO[subscript]2 concentration were measured. All plants were exposed to a drought treatment and were monitored daily until stomatal conductance was zero. At this point, critical leaf water potential (Ψ[subscript]crit), an indicator of physiological drought tolerance was assessed. Other measurements include root length, diameter, volume, and mass, leaf area, leaf tissue density, root tissue density, and root to shoot ratio. Traits were compared using pair-wise bivariate analysis and principal component analysis (PCA). A dichotomy was found between dry-adapted plants with thin, dense leaves and roots, high leaf angle, and highly negative Ψ[subscript]crit and hydrophiles which have the opposite profile. A second axis offers more separation based on high photosynthetic rate, high conductance rate, and leaf angle, but fails to provide a distinction between C[subscript]3 and C[subscript]4 species. When tested independently, grasses and forbs both showed drought tolerance strategies similar to the primary analysis. Matching up these axes with long term abundance data suggests that species with drought tolerance traits have increased abundance on Konza, especially in upland habitats. However, traits that relate to drought tolerance mirror relationships with nutrient stress, confounding separation of low water versus low nutrient strategies. My results not only illustrate the utility of morphological and physiological plant traits in classifying drought responses across a range of species, but as functional traits in predicting both drought tolerance in individual species and relative abundance across environmental gradients of water availability.

Doctor of Philosophy / Department of Horticulture, Forestry, and Recreation Resources / Dale J. Bremer / Jack D. Fry / Researchers and practitioners can use numerous techniques to measure or estimate evapotranspiration (ET) from turfgrass but little is known about how they compare to ET using standard lysimeters. An investigation was conducted to compare measurements of ET from lysimeters (LYS[subscript]E[subscript]T) with ET estimates from the FAO56 Penman-Monteith (PM[subscript]E[subscript]T) and Priestley-Taylor (PT[subscript]E[subscript]T) empirical models, atmometers (AT[subscript]E[subscript]T), eddy covariance (EC[subscript]E[subscript]T), and a canopy stomatal conductance model that estimates transpiration (COND[subscript]T). Methods were compared at the same site during the 2010, 2011, and 2012 growing seasons. Overall, PT[subscript]E[subscript]T and EC[subscript]E[subscript]T were not different from LYS[subscript]E[subscript]T, whereas PM[subscript]E[subscript]T, AT[subscript]E[subscript]T, and COND[subscript]T, increasingly underestimated LYS[subscript]E[subscript]T. Differences exist among ET measurement techniques and one should employ the technique that best fits their situation. An atmometer is an inexpensive tool that can be used to measure turfgrass ET within microclimates, such as those typically found in an urban home lawn. An investigation was conducted to compare AT[subscript]E[subscript]T estimates with PM[subscript]E[subscript]T estimates within a number of lawn microclimates. Home lawns in Manhattan and Wichita, KS, were selected for study during the growing seasons of 2010 and 2011. Open sward AT[subscript]E[subscript]T was 4.73 mm d[superscript]-[superscript]1, whereas PM[subscript]E[subscript]T was 5.48 mm d[superscript]-[superscript]1. Within microclimates, AT[subscript]E[subscript]T was 3.94 mm d[superscript]-[superscript]1 and PM[subscript]E[subscript]T 3.23 mm d[superscript]-[superscript]1. Atmometers can provide practitioners with reliable estimates of PM[subscript]E[subscript]T within microclimates. Zoysiagrass (Zoysia spp.) is a common turfgrass used on home lawns and golf courses. However, poor shade tolerance and cold hardiness have limited its use in the transition zone. A study was conducted to determine changes and differences in growth and physiology among selected Zoysia over a three-year period (2010-2012) in the transition zone. The genotypes were 'Emerald' [Z. japonica × Z. pacifica], 'Zorro' [Z. matrella], 'Meyer' and Chinese Common [Z. japonica], and experimental progeny Exp1 [Z. matrella × Z. japonica], and Exp2 and Exp3 [(Z. japonica × Z. pacifica) × Z. japonica]. 'Zorro' and 'Emerald' experienced winter injury. 'Meyer', Chinese Common, and Exp1 showed poor performance over the three-years. The Exp2 and Exp3 progeny, maintained high percent cover, visual quality, and tiller density, and may provide practitioners more shade-tolerant cultivar choices in the transition zone.

Master of Science / Agronomy / Mithila Jugulam / Common waterhemp (Amaranthus rudis) is a problematic weed species of cropping systems throughout the Midwestern states, including Kansas. Recently, waterhemp populations from Kansas were found to have evolved resistance to the widely used herbicide glyphosate as a result of amplification of the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), the enzyme target of glyphosate. The objectives of this research were to 1) perform glyphosate dose-response study and determine the relationship between relative EPSPS genomic copies and EPSPS gene expression in glyphosate-resistant waterhemp, and 2) characterize the genomic configuration and distribution of EPSPS copies using florescence in situ hybridization (FISH) in three glyphosate-resistant waterhemp populations. Waterhemp populations from eastern Kansas were screened with 868 g ae haˉ¹ (field used rate) of glyphosate, and genomic DNA and total RNA was isolated from the survivors to determine the EPSPS genomic copies and EPSPS gene expression relative to the acetolactate synthase (ALS) gene using qPCR. Furthermore, waterhemp specific EPSPS probes were synthesized to perform florescence in situ hybridization (FISH) on these glyphosate-resistant plants. Results of these experiments indicate a positive correlation between level of glyphosate resistance, EPSPS copies, and their expression. As expected, a negative correlation was found between shikimate accumulation and EPSPS copies. Sequencing of the EPSPS gene showed no presence of the proline 106 mutation, which is known to be associated with glyphosate resistance suggesting that an insensitive EPSPS enzyme was not involved in the mechanism of glyphosate resistance. FISH analysis of resistant plants illustrated presence of amplified EPSPS copies on two homologous chromosomes, likely near the centromeric region. . This is the first report demonstrating a positive relationship between EPSPS copies and expressions, as well as chromosome configuration of EPSPS copies in glyphosate- resistant waterhemp from Kansas.