Master of Science / Department of Entomology / Jeremy L. Marshall / Sexual conflict is a process that occurs when the evolutionary interests of the sexes is not aligned. The theory began with Darwin in the 1800’s. His observations of sexual dimorphism and traits which did not seem to follow the rules of natural selection led him to the theory of sexual selection. Sexual selection resolved some of the issues which were noted in the evolutionary processes he observed but not all of them. For example, it did not explain traits which increased the male optimum fitness while decreasing the female optimum fitness. It was decades before the concept of sexual conflict was formulated, and even longer before the underlying mechanisms were understood. Sexual conflict is different than sexual selection and there are ways to differentiate which of these processes have occurred in a population. The main forms of sexual conflict are intralocus and interlocus conflict. These involve the interactions between alleles in the genome of the sexes. Intralocus conflict involves conflict at a single locus in the genome while interlocus conflict involves conflict between different loci. Interlocus conflict has been more extensively studied due to its association with sexually antagonistic coevolution (SAC). SAC draws the attention of scientists due to the possibility it is related to the fundamental biological process of speciation. Sexual conflict targets certain traits at defined periods in the mating process. These periods include first, the precopulatory stage which is before the act of copulation begins. Second, there is the stage during copulation but before fertilization of the embryo. Finally, there is the postcopulatory postzygotic stage which is after copulation has ended and fertilization has occurred. Each of these points in the process of mating has traits or behaviors which sexual conflict may target. This review concludes with a proposed experiment to determine if sexual conflict is occurring in a group of four genera of mosquitoes. The experiment utilizes the attributes of sexual conflict to differentiate between other processes. A major component is the consequence of mating systems on selective processes to determine if sexual conflict is involved in the evolution of male accessory gland proteins.

Doctor of Philosophy / Department of Biology / Alexander E. Beeser / Multiple signaling pathways regulate development of the posterior zebrafish body, which is derived from a population of progenitor cells called the tailbud, a structure formed at the end of gastrulation. Fate specification and differentiation are closely linked with cell migration to ensure that, as some cells exit the tailbud and differentiate, other cells are retained in the tailbud as undifferentiated precursors to support later growth. The role of BMP signaling in specifying cell fate in the tailbud has been well-characterized. Among the lost ventral tissues like ventral tailfin and cloaca, embryos with compromised BMP signaling produce a curious phenotype-a ventrally located secondary tail containing both somitic muscle and notochord. This phenotype is proposed to be a fate-patterning defect when the BMP gradient lowered to a precise level. However, this morphogen mode is insufficient to explain secondary tail formation without considering BMP also regulates morphogenetic movements during gastrulation, promoting the convergence of lateral mesodermal cells towards the dorsal axis. In this study, we provide evidence that BMP signaling continues to mediate cell movements during tail development. Our data indicate that BMP signaling is activated in the ventroposterior tailbud to promote cell migration during tailbud protrusion, and that it is the defective migration of these cells which ultimately leads to bifurcation of the CNH domain, a presumptive stem cell pool in the tailbud, and formation of a secondary tail in BMP mutants. In parallel, the morphogenesis of tailbud cells is known to be under the control of noncanonical Wnt signaling, although the exact nature of the defect remains unclear. We find that inhibition of noncanonical Wnt signaling also leads to secondary tail formation. Additionally, we show that noncanonical Wnt signaling interacts with BMP signaling to maintain CNH integrity by affecting cadherin localization in CNH cells, possibly disrupting cell cohesion. We propose a model that BMP and a noncanonical Wnt pathway regulate tail morphogenesis by controlling cell migration and cell adhesion within the tailbud.

Doctor of Philosophy / Genetics Interdepartmental Program / Bikram S. Gill / Threshability is an important agronomic trait in wheat as free-threshing forms facilitate mechanical threshing of grain. All wild relatives of wheat have tough glumes and are non-free-threshing, whereas most cultivated wheats have soft glumes and are free-threshing. Two genetic loci are known to govern the threshability trait in bread wheat. The Q gene located on chromosome 5AL and glume tenacity genes located on homoeologous group-2 chromosomes seem to interact to produce a free-threshing phenotype. Although, the Q gene was found to be a member of APETALLA 2 (AP2) class of transcription factors, the molecular nature of the tough glume genes remains unknown. In the present study, genetic and molecular characterization of two of the threshability genes in wheat was undertaken. The soft glume (sog) gene of diploid wheat and tenacious glume (Tg) gene of hexaploid wheat were characterized and mapped on short arm of chromosome 2Am and 2D respectively. Comparative mapping of sog and Tg genes suggested their independent origins. The sog gene was mapped in a low-recombination region near the centromere on 2AmS. Genomic targeting using deletion bin mapped ESTs assigned the Tg gene to a 4.9 cM interval in the distal 16% of short arm of chromosome 2D. In order to find additional markers for fine-mapping the Tg gene, macrocolinearity between rice and wheat was explored in the Tg region. Although synteny between rice and wheat was found to be conserved in the distal region of chromosome 2DS, the genomic region encompassing the Tg gene in wheat showed some rearrangements relative to rice. Molecular characterization of ethyl methanesulfonate-induced free-threshing mutants in two different non-free-threshing backgrounds revealed point mutations as well as variable sized deletions at Tg locus. Targeting of Tg to the high-recombination gene-rich region in wheat and availability of several genomic resources from the present study will aid in the cloning and further characterization of this important agronomic gene.

Master of Science / Department of Statistics / Gary L. Gadbury / QTL (quantitative-trait locus) analysis aims to locate and estimate the effects of genes that are responsible for quantitative traits, such as grain protein content and yield, by means of statistical methods that evaluate the association of genetic variation with trait (phenotypic) variation. Quantitative traits are typically polygenic, i.e., controlled by multiple genes, with varying degrees of in uence on the phenotype. Several methods have been developed to increase the accuracy of QTL location and effect estimates. One of them, multiple interval mapping (MIM) (Kao et al. 1999), has been shown to be more accurate than conventional methods such as composite interval mapping (CIM) (Zeng 1994). Other QTL analysis methods have been developed to perform additional analyses that might be useful for breeders, such as of pleiotropy and QTL-by-environment (QxE) interaction. It has been shown (Jiang and Zeng 1995) that these analyses can be carried out with a multivariate extension of CIM (MT-CIM) that exploits the correlation structure in a set of traits. In doing so, this method also improves the accuracy of QTL location detection. This thesis describes the multivariate extension of MIM (MT-MIM) using ideas from MT-CIM. The development of additional multivariate tests, such as of pleiotropy and QxE interaction, and several methods pertinent to the development of MT-MIM are also described. A small simulation study shows that MT-MIM is more accurate than MT-CIM and univariate MIM. Results for real data show that MT-MIM is able to provide a more accurate and precise estimate of QTL location.

Doctor of Philosophy / Department of Plant Pathology / Bikram S. Gill / Segregation distortion describes the transmission of an allele or alleles of a heterozygous locus at a higher frequency than expected in a Mendelian ratio. From the organism's view, segregation distortion is the preferential retention of chromosomal blocks carrying genes beneficial to its fitness and reproductive viability. In wheat the best studied segregation distortes are those introduced from Aegilops species; these selfish genetic elements are named gametocidal (Gc) genes and the chromosomes carrying them are called Gc chromosomes. This genetic mechanism causes chromosome breakage in gametophytes lacking the Gc carrier chromosome, thus favoring its own retention in the genome. While the mode of action of the Gc genes is not yet known, they have been used extensively in wheat genetics for the development of deletion stocks, a key resource for elucidating the structure of physical regions containing important genes. The objective of this study was to develop the tools necessary to map the Gc2 gene derived from Ae. sharonensis and perform map-based cloning. Extensive physical and genetic mapping located the gametocidal gene on the distal 1% of the 4BL arm present in the T4BS[dot in middle of line]4BL-4S[sh superscript]#1L translocation chromosome. Comparative genomics using rice provided markers distal and proximal to the Gc2 locus; however, synteny broke down at the locus. The characterization of this chromosomal region has provided insight into its recombination frequency, synteny and composition; however, the dynamic architecture of the end of the chromosome has made comparative mapping of this region difficult.

Doctor of Philosophy / Department of Animal Sciences and Industry / David M. Grieger / Early in embryonic development a series of events occur whereby pluripotent cells undergo differentiation to give rise to the three germ layers and extraembryonic tissues of the developing conceptus. Nanog, Sox-2, and Oct-4 genes have been identified as having key roles in maintaining pluripotency in undifferentiated human and mouse cells but recent evidence suggests they may have different roles in farm animals. We cloned the coding sequence for porcine Nanog including 452 base pairs of the Nanog promoter, and partial coding sequences of Oct-4 and Sox-2. Embryos were flushed from sows 10, 12, 15, and 17 days post insemination. RNA was isolated from whole d-10 and -12 conceptuses, d-15 embryonic disk, distal and proximal extraembryonic tissue, and d-17 embryonic disk, distal and proximal extraembryonic tissue, and allantois for real-time PCR. RNA from d-40 maternal myometrium and endometrium, fetal placenta, and liver were also used in real-time PCR. The homeodomain and c-terminal tryptophan repeats are highly conserved in porcine Nanog compared to the mouse, human and bovine. In the promoter, the highly conserved Octamer and Sox binding sequences are also present. The Nanog expression pattern was different when compared to Oct-4 and Sox-2. Day-40 tissues demonstrated the highest expression including endometrium (7 fold) fetal liver (27 fold), placenta (40 fold) and myometrium (72 fold) when compared to day 15 distal extraembryonic tissue. Oct-4 and Sox-2 expression was lowest in d-40 tissues except for fetal liver which was 20 and 71 fold, respectively, higher than endometrium. Oct-4 levels were consistent in d-10, -12, and -15 conceptuses and disk but dropped 3 fold in d-17 disk. On the other hand, Sox-2 was upregulated a 1000 fold in the d-15 disk and 2000 fold in the d-17 disk when compared to the d-12 conceptus. Nanog may have other roles in than maintenance of pluripotency including a possible role in multipotent or progenitor stem cells. Expression of all 3 markers in fetal liver suggests a more primitive cell type is present such as hematopoietic stem cells.

Master of Science / Department of Agronomy / John P. Fellers / Allan K. Fritz / Water availability is commonly the most limiting factor to crop production, especially in drought prone areas like the Midwest. This study was conducted to map quantitative trait loci (QTL) involved in drought tolerance in wheat (Triticum aestivum L.) to enable their use for marker assisted selection (MAS) in breeding. A population of 122 F[subscript]7 derived recombinant inbred lines from a cross between Dharwar Dry and Sitta, spring wheat lines with contrasting drought tolerances, was analyzed using the amplified fragment length polymorphism (AFLP) technique and Diversity Array Technology (DArT) markers to create a QTL map. Of the 256 AFLP primer combinations evaluated, 151 were found to be polymorphic between the parents and were used to screen the population. A linkage map of 48 groups was created from the combined DArT markers, AFLP data, and SSR markers. This was used to create a QTL map which identified QTL in 24 of these groups. Using these markers for MAS in a breeding program could overcome the difficulties of selecting for drought tolerance. Another serious limitation to wheat production is leaf rust caused by the pathogen Puccinia triticina. Leaf rust causes between 1% and 20% yield loss on average and tends to be the worst in years with high yield potential. PI 289824 contains a single, dominant gene for seedling resistance mapping to chromosome 5BS and thought to be different from Lr52. An F[subscript]2 mapping population from a cross between PI 289824 and Jagger was used to try to identify markers very closely linked to the gene and therefore useful for MAS. The population presented some mapping challenges, but with the use of SSR and EST-STS markers, the gene was flanked. However, the markers were at too a great distance to be useful for mapping.

Doctor of Philosophy / Genetics Interdepartmental Program-Agronomy / William T. Schapaugh Jr / The soybean aphid (Aphis glycines Matsumura) has been one of the major pests of soybean [Glycine max (L.) Merr.] in soybean-growing regions of North America since it was first reported in 2000. The objectives of this study were to screen for soybean aphid resistant genotypes, determine the inheritance of resistant genes, and map and validate the resistance gene in the moderate resistant genotype K1621 using simple sequence repeat (SSR) markers. A mapping population of 150 F2:3 families from the cross between K1621 and susceptible genotype KS4202 were evaluated for aphid resistance. Phenotyping was conducted on the basis of total aphid number per plant 7 days following infestation with 4 aphids. Inheritance study indicated that one major dominant gene controls soybean aphid resistance in K1621. After SSR markers for polymorphism were screened between parents, a total of 133 polymorphic markers distributed across the soybean genome were used for genotyping. One quantitative trait loci (QTL) controlling antibiotic resistance was found by using the composite interval mapping method. This QTL localized on chromosome 13 (linkage group F) between markers Sat_234 and S6814 and explained 54% of the phenotypic variation. The putative QTL was further validated by single marker analysis using an independent population derived from the cross of K1621 and Dowling. The locus for soybean aphid resistance in K1621 was named [Rag]_K1621. The markers identified and validated in this study could be useful for marker-assisted selection of [Rag]_K1621.

Doctor of Philosophy / Department of Agronomy / Allan K. Fritz / In this research, a recombinant inbred line (RIL) population derived from cross between a leaf rust- and stripe rust-susceptible spring wheat ‘Avocet S’ and a slow leaf- and stripe-rusting resistant spring wheat ‘Amadina’ was used to postulate and map leaf rust seedling resistance genes, identify quantitative trait loci (QTL) for slow-rusting resistance against leaf and stripe rust, and study slow leaf-rusting components, latent period and infection frequency. Two known Lr genes (Lr23, and Lr26) were identified to be present in ‘Amadina’ through gene postulation, pedigree, cytogenetic, and polymerase chain reaction analyses. One unknown gene associated with seedling resistance was also mapped on chromosome 1BL. In greenhouse experiment, it was estimated that at least five genes conditioning final disease severity (FS) and latent period (LP), and four genes conditioning infection frequency (IF), segregated in the population. Correlations between LP and FS, and LP and IF were moderately negative, and that between IF and FS was moderately positive, indicating inter-dependence of the traits. Two QTL on chromosomes 1BL and 6BL were associated with LP and FS, and three QTL on chromosomes 1BL, 6BL and 2DS were associated with IF. Segregation of the RIL population in field experiment indicated that there were at least four and three adult plant resistance (APR) genes involved in resistance for leaf and stripe rust. Six QTL on chromosomes 3AL, 4AL, 1BL, 5BL, and 7BL were associated with APR for leaf rust, and seven QTL on chromosome 4AL, 5AL, 1BL, 2BL, 4BL, 5BL, 2DL, and 4D were associated with APR for stripe rust. Our results indicated that the major portion of genetic variability for slow-rusting resistance was additive gene action, and, to some extent, epistasis. In this research, we also explored the utility of remote sensing and geographic information systems (GIS) and analytical operations to discriminate leaf rust pustules from other parts of leaf and to accurately determine pustule size in ‘Amadina’ and ‘Avocet S’.

Doctor of Philosophy / Department of Biology / Jyoti Shah / Localized inoculation of a plant with an avirulent pathogen results in the activation of systemic acquired resistance (SAR), a defense mechanism that confers enhanced resistance against a variety of pathogens. The activation of SAR requires the translocation of an unknown signal from the pathogen-inoculated organ to the other organs where defenses are primed to respond faster in response to a future attack by a pathogen. Previous studies with the Arabidopsis thaliana dir1 (defective in induced resistance1) and sfd1 (suppressor of fatty acid desaturase deficiency1) mutants implicated a role for plant lipids in the activation of SAR. DIR1 encodes a putative lipid transfer protein and SFD1 encodes a dihydroxyacetone phosphate (DHAP) reductase involved in plastid glycerolipid metabolism. To further evaluate the role of DHAP reductases and plastid lipids in SAR, the involvement of two additional putative DHAP reductase encoding genes (AtGPDHp and AtGPDHc) and the SFD2 gene, which like SFD1 is involved in plastid glycerolipid metabolism, in SAR was evaluated. Only SFD2 was found to be essential for SAR. Although the lipid profile of the sfd2 mutant was similar to that of the fad5 (fatty acid desaturase 5) mutant, sfd2 is not allelic with fad5 and does not influence FAD5 expression. The SFD2 gene was mapped to an 85 kilo basepairs (kb) region on the third chromosome of Arabidopsis. The lipid composition defect of the sfd2 mutant was partially complemented by two independent recombinant bacterial artificial chromosomes (BACs) that contained genomic DNA spanning the wild type SFD2 locus. The role of plastid synthesized glycerolipids in the activation of SAR was further evaluated by characterizing SAR in additional Arabidopsis mutants that were deficient in plastid lipid metabolism. The requirement of MGD1 (MONOGALACTOSYLDIACYLGLYCEROL SYNTHASE 1), DGD1 (DIGALACTOSYL-DIACYLGLYCEROL SYNTHASE 1) and FAD7 (FATTY ACID DESATURASE 7) genes in SAR, confirmed the essential role of plastid glycerolipids, presumably a galactolipid-dependent factor, in signaling associated with the SAR.