Doctor of Philosophy / Department of Agronomy / Allan K. Fritz / Heat stress is one of the major environmental constraints for wheat production worldwide. High temperature during grain filling in wheat leads to a significant reduction in yield. In this research, three different projects were completed. The first project was to study cytoplasmic effects on heat tolerance in wheat, where ten different alloplasmic lines of wheat were backcrossed with four different wheat varieties: „Karl 92‟, „Ventnor‟, „U1275‟ and „Jagger‟. The nuclear genome of the alloplasmic lines was substituted by backcrossing six times using the recurrent parents as males. During the fifth and/or fourth backcross, reciprocal crosses were made to develop NILs (Near Isogenic Lines) for cytoplasm. Sixty-eight NILs and their parents were evaluated in growth chambers for post-anthesis heat tolerance. Plants were grown in the greenhouse and placed under heat stress for 14 days starting at 10 days after anthesis. Growth chambers were maintained at 35°/30°C for heat stress and the greenhouse was maintained at 20°/15°C as the optimum temperature. Effects of high temperature on chlorophyll content and Fv/Fm (a chlorophyll fluorescence measuring parameter) were found to be significant. Cytoplasms 1, 4, 5, 8, 9 and10 provided greater tolerance in one or more nuclear backgrounds. These results indicated that cytoplasmic effects can contribute to heat tolerance of wheat. The second project focused on identification of quantitative trait loci (QTL) for thylakoid membrane damage (TMD), SPAD chlorophyll content (SCC) and plasma membrane damage (PMD), as these traits are found to be associated with resistance to heat stress and contributes to relatively stable yield under high temperature. A RIL (Recombinant Inbred line) population of a cross between winter wheat cultivars „Ventnor‟ and „Karl 92‟ was evaluated using two different temperature regimes (20°/15°C, 36°/30°C) imposed at ten days after anthesis. The aforementioned traits were evaluated and associated with various molecular markers (SSR, AFLP and SNP). The putative QTL associated are localized on chromosomes 6A, 7A, 1B, 2B and 1D and have the potential to be used in marker assisted selection for improving heat tolerance in wheat. In the third project, a transgenic approach to increase grain fill during high temperatures was investigated. Grain fill is reduced at temperatures above 25°C in wheat partly due to the inactivity of soluble starch synthase. We isolated a soluble starch synthase gene from rice that has the potential to overcome this deficiency during high temperatures and placed it behind both a constitutive promoter and an endosperm-specific promoter. Transgene expression and the effects of the transgene expression on grain yield-related traits for four generations (T0, T1, T2 and T3) were monitored. The results demonstrated that even after four generations, the transgene was still expressed at high levels, and transgenic plants produced grains of greater seed weight than Bobwhite control plants under the same environmental conditions. Thousand-seed weight under high temperatures increased 21-34% in T2 and T3 transgenic plants when compared to the non-transgenic control plants. In addition, the duration of photosynthesis was longer in transgenic wheat than in non-transgenic controls. Our study demonstrated that expression of rice soluble starch synthase gene in wheat can improve wheat yield under heat stress conditions.

Master of Science / Department of Horticulture, Forestry, and Recreation Resources / Jack Fry / Zoysiagrass (Zoysia spp.) is more heat and drought resistant and requires fewer cultural inputs than cool-season (C3) turfgrasses; however, its widespread use may be limited by an extended period of winter dormancy, the disease Rhizoctonia large patch (Rhizoctonia solani Kühn AG-2-2 LP), and lack of cold hardiness. Objectives of these 2012-2014 field studies were to evaluate: 1) turfgrass colorants and overseeding for color enhancement; 2) three colorant application volumes on color persistence; 3) colorants applied at two volumes, once or sequentially, on buffalograss (Buchloë dactyloides) and zoysiagrass; 4) the impact of nitrogen source and application timing on large patch severity; and 5) winter hardiness and turf quality of new experimental lines. The colorants Ultradwarf Super and Green Lawnger provided acceptable lawn-height ‘Chisholm’ zoysiagrass color for 7 to 9 and 19 to 24 weeks after treatment (WAT), respectively. Chisholm receiving a sequential application of either product in mid-winter (14 WAT) maintained an acceptable color level after that point until mid-May (28 WAT). Overseeding with annual ryegrass did not provide acceptable color for more than 4 weeks. Evaluation of the colorants Green Lawnger, Endurant, and Wintergreen Plus showed that acceptable Chisholm color at lawn-height occurred for 8 to 14 WAT at 80 gallons per acre (GPA) and 16 to 26 WAT at 240 GPA. Buffalograss at lawn-height receiving a single autumn colorant application had acceptable color for 8 to 12 WAT at 100 GPA or 8 to 14 WAT at 160 GPA. ‘Meyer’ zoysiagrass maintained at a 0.5 inch height receiving a single autumn colorant application had acceptable color for 8 to 18 WAT at 100 GPA or 14 to 18 WAT at 160 GPA. No differences in large patch occurred between spring and fall applications of ammonium sulfate and calcium nitrate, nor between those treatments and summer-applied urea. Applying fertilizer in spring when soils reached 21 ºC increased green cover on some rating dates compared to applications in fall when soil temperatures fell to 21 ºC but differences were minimal. Out of 985 experimental zoysiagrass lines planted in the field, about 25 were identified for further evaluation for cold hardiness, large patch resistance, and turf quality.

Master of Science / Department of Horticulture, Forestry, and Recreation Resources / Kimberly A. Williams / Ivy geranium (Pelargonium peltatum L’Herr ex. Ait.) and tomato (Solanum lycopersicum L.) are two economically important greenhouse crops known to be affected by non-pathogenic lesions on leaf tissues. These physiological disorders are often termed edema (oedema) or intumescences, but several other names have been used including enations, non-pathogenic galls or tumors, and neoplasms. These lesions, characterized by small protrusions on leaf tissues that become necrotic over time, are considered to be the result of environmental factors. Our research focused on determining what environmental factors affect these disorders on ivy geranium and tomato. The physiological disorder of ivy geranium is thought to be the result of water uptake exceeding transpiration, resulting in a build-up of water and solutes in leaf tissue that results in the blister-like protrusions in the epidermal layer. Current convention suggests that susceptible plants be grown in an environment that promotes transpiration with low humidity and infrequent watering. Over four experiments, we evaluated the effects of four root medium water contents, five rates of supplemental calcium application and two vapor pressure deficit (VPD) environments on three cultivars of ivy geranium. Our results indicate that high root medium water contents do not increase the incidence of edema on ivy geranium, but increase overall plant growth. Supplemental calcium had no affect on edema or growth, while our VPD results were inconclusive. These results suggest that current convention regarding cultural practices that abate the disorder be revisited. In tomato var. hirsutum ‘Maxifort’, the physiological disorder is characterized by individual epidermal cells swelling, which is unlike the disorder in ivy geranium where solutes build up across a group of epidermal cells. The environmental factors we focused on were two root medium water contents and supplemental UVB light. Our results suggest that root medium water content may play a role in development of tomato intumescences based on visual observation, and UVB light supplementation helps prevent the lesions from forming.

Master of Science / Department of Agronomy / Rob M. Aiken / Mary Beth Kirkham / Prior studies indicate clumped planting can increase grain sorghum yield up to 45% under water deficit conditions by reducing tiller number, increasing radiation use efficiency, and preserving soil water for grain fill. The objective of this study was to evaluate effects of planting geometry on sorghum grain yield. The field study was conducted in seven environments with two sorghum hybrids, four populations, and two planting geometries. Crop responses included leaf area index, yield, and components of yield. Delayed planting decreased yield by 39%, and a later maturing hybrid increased yield, relative to an early hybrid, by 11% under water sufficiency. Clumped planting increased the fraction of fertile culms (culms which formed panicles) from 5-14%. It reduced the number of culms m-2 by 12% under water limiting conditions (at one of two locations) but increased culms m-2 16% under water sufficiency. Seeds per panicle and seed weight generally compensated for differences in panicles m-2, which were related to different planting population densities. Although agronomic characteristics of hybrids varying in maturity have been widely studied, little information exists concerning their physiological differences. Therefore, the objective of the greenhouse study was to determine if stomatal resistance, leaf temperature, and leaf chlorophyll content differed between two DeKalb grain sorghum [Sorghum bicolor (L.) Moench] hybrids. They were DKS 36-16 and DKS 44-20, of medium-early and medium maturity, respectively, when grown under field conditions in Kansas. Seeds were planted in a greenhouse. Stomatal resistance and leaf temperature were measured 55 days after planting with a Decagon Devices (Pullman, WA) diffusion porometer, and chlorophyll content was measured 119 days after planting with a Konica Minolta (Osaka, Japan) SPAD chlorophyll meter. The two hybrids did not differ in stomatal resistance, leaf temperature, chlorophyll content, height, and dry weight. Their difference in maturity was not evident under the greenhouse conditions. Future work needs to show if hybrids of different maturities vary in physiological characteristics

Master of Science / Department of Plant Pathology / Harold Trick / The object of this study was to find a target sequence for the known Heterodera glycines Y25 sequence that contained no homology to any known Glycine max genes so homologous endogenous soybean gene expression will not be effected. In addition, in attempt to improve the accuracy of SCN bioassays performed in greenhouse settings, applications of a variety of insecticides with differing modes of action were applied to screen for any detectable effects on the SCN populations. The full-length sequence of the Y25 gene was blasted against the G. max genome using the National Center for Biotechnology Information blast database and a portion of the gene was found to contained no homology to the G. max genome. A rapid hairy root assay was used to screen for resistance to H. glycines. The sequence was transformed into Agrobacterium rhizogenes using a modified heat shock method. The transformed A. rhizogenes were used to inoculate soybean seedlings. The inoculated seedlings developed hairy roots expressing the target sequence. Upon finishing the hairy root assay it was discovered that there were no detectable differences across any of the treatments or the controls. It was neither proved nor disproved that the new target sequence containing no homology to the G. max genome was as effective as the original target. Further investigation will need to be conducted to show the level of control for the new target sequence.

Doctor of Philosophy / Department of Horticulture, Forestry, and Recreation Resources / Jack Fry and Megan Kennelly / Rough bluegrass (RBG, Poa trivialis L.) is a difficult-to-control weed that commonly infests cool-season turfgrass swards after movement of vegetative propagules or contamination from seed lots. Rough bluegrass is less tolerant of heat stress than desirable cool-season species such as tall fescue (TF, Festuca arundinacea Schreb. Syn [italicize]Schedonorus [italicize]arundinaceus Schreb.), and often declines during mid-summer due to biotic or abiotic stresses. The objectives of these 2011-2013 controlled environment and field experiments were to: 1) observe growth and physiological differences between ‘Laser’ and ‘Pulsar’ RBG and TF; 2) differentiate between physiological and pathological contributors to RBG decline; 3) determine the effects of TF seeding rate and mowing height on TF/RBG establishment when RBG is a seed contaminant; 4) evaluate herbicide combinations for selective RBG control; and 5) evaluate seasonal timing of glyphosate for nonselective RBG control. Tall fescue was less affected by elevated temperature than RBG. At 35°C, Laser and Pulsar experienced similar reductions in quality, gross photosynthesis (Pg), shoot and root biomass, and root length density compared to when grown at 23°C, but maximum electrolyte leakage was greater for Pulsar (63%) than for Laser (49%). Cell membrane thermostability could contribute to the better heat tolerance of Laser RBG. Evaluation of RBG foliage and roots did not reveal a fungal pathogen associated with RBG decline. Still, repeated applications of azoxystrobin (610 g a.i. ha⁻¹) or pyraclostrobin (556 g a.i. ha⁻¹) increased RBG quality, cover, and Pg during summer compared to untreated RBG, possibly due to poorly understood non-target physiological effects of the fungicides. Mowing TF at 7.6 or 11.4 cm reduced RBG incidence up to 57% compared to mowing at 3.8 cm. Tall fescue seeding rate had no effect on RBG incidence. Several herbicides and herbicide combinations resulted in some RBG injury in the field, but bispyribac-sodium was the only treatment that provided RBG control (16 to 92%) in Manhattan, KS; Hutchinson, KS; and Mead, NE. Spring-applied glyphosate resulted in the lowest RBG coverage (1 to 31%) among field studies in Manhattan and Mead, followed by late-summer applications (6 to 58%), and mid-summer applications (9 to 86%).

Master of Science / Department of Agronomy / William T. Schapaugh / Heat and drought stress are common problems for crops grown in Kansas. Rarely do these problems occur separately, more often than not they occur in tandem if not simultaneously. The interaction of heat stress and pollen germination was investigated in order to determine if a physiological screen was a feasible method of determining heat tolerance in soybean [Glycine max (L.) Merr]. Ten soybean lines (Group A) from the 2006 Northern Region Uniform Soybean Tests were analyzed over two years in four locations consisting of irrigated and dryland field environments, with an additional twenty lines (Group B) analyzed in the second year. Pollen was collected from plants and incubated at either 28o, 34o, or 38o C to determine pollen germination for optimal and stress-inducing temperatures. A three-way interaction of entry x incubation temperature x environment was observed, as well as significant differences among entries, incubation temperatures and environments. Average pollen germination for soybean entries ranged from 25% to 38% across three incubation temperatures and four environments in Kansas during 2006 – 07. The average environment effect for pollen germination ranged from 29% (dryland, 2006) to 34% (irrigated, 2007), while the average incubation temperature effect on pollen germination ranged from 25% (38oC) to 44% (28oC). This experiment has shown that increasing incubation temperatures significantly decreases pollen germination in vitro. It has also shown that soybean genotypes differ in pollen germination and that an in vitro screen can be used to characterize these differences. Further studies are needed to establish the relationship between pollen germination, seed set and seed yield in soybean. Work also needs to be completed to determine the proper sample size to adequately characterize differences in pollen germination so that performance differences among genotypes can be used as selection criteria in a plant breeding program.

Master of Science / Department of Agronomy / William T. Schapaugh Jr / Crops research moving forward faces many challenges to improve crop performance. In breeding programs, phenotyping has time and economic constraints requiring new phenotyping techniques to be developed to improve selection efficiency and increase germplasm entering the pipeline. The objectives of these studies were to examine the changes in spectral reflectance with soybean breeding from 1923 to 2010, evaluate band regions most significantly contributing to yield estimation, evaluate spectral reflectance data for yield estimation modeling across environments and growth stages and to evaluate the usefulness of spectral data as an optimized phenotyping technique in breeding programs. Twenty maturity group III (MGIII) and twenty maturity group IV (MGIV) soybeans, arranged in a randomized complete block design, were grown in Manhattan, KS in 2011 and 2012. Spectral reflectance data were collected over the growing season in a total of six irrigated and water- stressed environments. Partial least squares and multiple linear regression were used for spectral variable selection and yield estimation model building. Significant differences were found between genotypes for yield and spectral reflectance data, with the visible (VI) having greater differences between genotypes than the near-infrared (NIR). This study found significant correlations with year of release (YOR) in the VI and NIR portions of the spectra, with newer released cultivars tending to have lower reflectance in the VI and high reflectance in the NIR. Spectral reflectance data accounted for a large portion of variability for seed yield between genotypes using the red edge and NIR portions of the spectra. Irrigated environments tended to explain a larger portion of seed yield variability than water-stressed environments. Growth stages most useful for yield estimation was highly dependent upon the environment as well as maturity group. This study found that spectral reflectance data is a good candidate for exploration into optimized phenotyping techniques and with further research and validation datasets, may be a suitable indirect selection technique for breeding programs.

Master of Science / Genetics Interdepartmental Program / Bikram S. Gill / TILLING (Targeting Induced Local Lesions IN Genomes) is a high-throughput, reverse genetics strategy for scanning mutagenized populations for point mutations in loci of interest. Originally, TILLING was used to investigate gene function in Arabidopsis and has since been similarly applied for gene functional analysis in other organisms. TILLING also allows the generation of novel genetic variation in specific genotypes and, thus, has been implemented as a tool for crop improvement. Ethyl methanesulfonate (EMS) is a widely used mutagen to induce point mutations in most TILLING protocols. M1 plants are then self-pollinated and M2 seed harvested. A single seed is grown from each M2 progeny and tissue taken for DNA isolation. M3 seed is cataloged. DNA is pooled to increase the efficiency and aid in mutation detection. Polymerase chain reaction (PCR) is used to amplify a locus of interest using the M2 DNA pools as a template. The PCR products are digested with an endonuclease that cleaves mismatched, mutant DNA, and the digested products are visualized. The pools for which PCR products are positive for a mutation are deconvoluted to determine which individual plant of the pool was responsible for the mutation. DNA from the positive individual is sequenced to determine the type of mutation (missense, nonsense, synonymous). Individuals with mutations that are more likely to disrupt gene function (nonsense and certain missense) are studied further by growing the corresponding M3 generation. In bread wheat, Triticum aestivum, TILLING is complicated by polyploidy: genes that have homoeologs require that the functionality of each be studied. If functional homoeologs are present for all three genomes, mutants must be identified for each homoeolog, followed by successive intercrossing to produce a triple mutant plant. As a model for wheat genetics, we propose TILLING in diploid wheat. EMS mutant populations were created in diploid wheat (Triticum monococcum ssp. monococcum) and the hexaploid bread wheat cultivar ‘Jagger’. The diploid and hexaploid wheat populations were screened for mutations at the waxy locus, GBSS1, as a validation of our population and for comparative analysis of mutation rates in 2x and 6x wheat. For diploid wheat, GBSSI was screened in 716 M2 plants, and one mutant was found for 1.9 Mb screened. 3 For hexaploid wheat, GBSSI was screened in 518 M2 plants, and 30 mutants were identified within a total of 657 Kb screened, giving a mutation frequency of one mutation per 22 Kb. The reasons for this vast difference in mutation frequency between diploid and hexaploid wheat are discussed. The diploid wheat population was further examined by screening for mutations within four lignin biosynthesis candidate genes, for a total of 2 Mb screened. A single mutant was discovered for both of the lignin genes PAL6 and HCT, giving a mutation frequency of one mutation per 1 Mb screened.

Doctor of Philosophy / Department of Agronomy / P.V. Vara Prasad / Sorghum (Sorghum bicolor L. Moench) is the fifth most economically important cereal crop grown worldwide and adapted to a wide range of climatic conditions. Drought stress has been ranked as one of the most significant causes of crop yield loss with its effects on yield and yield components. Conservative water use by plants is one of the strategies that can be used as a drought coping mechanism. The slow wilting trait has been associated with conservative water use and has been found in some sorghum genotypes. The purpose of this study was to use canopy temperature to screen for drought tolerance in sorghum, evaluate water use efficiency for slow wilting sorghum genotypes and determine variability in root morphology and response to drought among sorghum genotypes. Canopy temperature studies were conducted under field conditions using infrared (IR) sensors while water use efficiency and root studies were conducted under greenhouse conditions. Our results showed a distinct separation in canopy temperature among genotypes under field conditions at 2:00 pm to 6:00 pm. Midday canopy temperature depression (CTD) was positively correlated to yield (R2 = 0.19) and harvest index (R2 = 0.11). CTD was also stable for all the genotypes during the period from 1:00 pm to 7:00 pm. There was a negative correlation between CTD and crop water stress index (CWSI) (R2 = 0.34) and a positive one between canopy temperature and CWSI (R2 = 0.50). Evaluation of genotypes for water use efficiency revealed significant variability among sorghum genotypes in the amount of water used (10.48 – 13.52 kg) and transpiration efficiency (TE) (2.64 – 7.11 g kg-1) among genotypes. Slow wilting genotypes were high in TE. Rooting depth increased for some genotypes under drought stress with genotype SC1124 recording the largest increase (180%). Total root length for some genotypes increased by 11 – 113% with genotypes SC224 and SC1019 recording the greatest increase. There was a positive correlation between water used and root length (R2 = 0.21). These results show that there is potential for selection of drought tolerance in sorghum and that genotypes with the slow wilting traits are efficient in water use.