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 Agronomy / Guihua Bai / Stem rust, caused by Puccinia graminis f. sp. tritici, has successfully prevented rust epidemics by Deployment of resistant cultivars in the past several decades. Unfortunately, race TTKS (termed Ug99) has defeated most stem rust resistance genes existing in commercial cultivars. Sr40, a stem rust resistance gene from Triticum timopheevii ssp. araraticum, was transferred to wheat and provides effective levels of seedling and adult plant resistance against Ug99. To characterize Sr40 in wheat, two mapping populations were developed from the crosses RL6088 / Lakin and RL6088 / 2174. RL6088 is an Ug99-resistant parent with Sr40. Since race TTKS is a quarantined pathogen, a US stem rust isolate RKQQ that is avirulent to Sr40 was used to evaluate the rust resistance in the F[subscript]2 and F[subscript]2:3 populations at the seedling stage. A total of 83 simple sequence repeats (SSR) primers on chromosome 2B were used to screen the parents for polymorphism. Each F[subscript]2 population was analyzed with the markers polymorphic between two parents. Marker Xwmc344 was the most closely linked to Sr40, at 0.7 cM proximal, in the linkage map constructed from the population RL6088 / Lakin, while Xwmc474 and Xgwm374 were also tightly linked. Xwmc474 was mapped 2.5 cM proximal to Sr40 in the RL6088 / 2174 population. Xwmc474 and Xwmc661 were flanking markers for Sr40 in both populations. Markers linked to Sr40 will be useful for marker-assisted integration of Sr40 into elite wheat breeding lines. In addition, a unknown stem rust resistance gene from another source, OK01307, a breeding line from Oklahoma State University shows partial resistance to Ug99, and was characterized using SSRs in this study. Two mapping populations were developed from cross OK01307 / Chinese Spring and OK01307 / LMPG-6. A total of 1300 SSR primers were screened for polymorphism between OK01307 and Chinese spring, and 1000 SSR primers were screened for polymorphism between OK01307 and LMPG-6. Polymorphic primers between parents and between bulks were used to screen the corresponding population. One Sr gene in OK01307 was mapped on chromosome 1BS of the both populations, which was closely linked to Sr24. Whether the gene is Sr24 per se or a new Sr gene that closely linked to Sr24 needs further investigation.

No description available.

DAWDLE (DDL) is one of the eighteen genes in Arabidopsis thaliana that encodes a protein with a Fork-Head Associated domain, a phospho-threonine binding domain providing a role in DNA repair and cell cycle regulation. DDL also contains an arginine-rich N terminal domain with putative Nuclear Localization Signals and a region for RNA binding. Two ddl knockout alleles in the WS-2 ecotype exhibit a strong pleiotropic phenotype with developmental defects such as short root and hypocotyl, reduced fertility, and distorted organs. This developmentally delayed phenotype is due to reduced accumulation of microRNAs and the phenotype is similar to those displayed by mutants involved in microRNA biogenesis pathway, suggesting a function for DDL in the microRNA biogenesis. One of the goals of my research was to characterize DDL protein through a structureunction study. Twelve point mutants were isolated in a mutagenesis screen and a comparative phenotypic and molecular characterization of each mutant with wildtype (WT) plants was performed. This revealed a few functionally significant amino acid residues of DDL. Traits for comparison included hypocotyl and root length, plant height, fertility, and microRNA accumulation. While all the mutants displayed reduced fertility, some of them had significantly varying stem height, hypocotyl and root length, and microRNA accumulation compared to the WT. Another objective of my research was to identify components involved in the DDL pathway, which in-turn would contribute to discovering additional components in microRNA biogenesis pathway. One such component, ddl suppressor1 (dds1), was identified through a second site mutagenesis screen of ddl-2. Phenotypic and molecular characterization revealed that dds1 is a strong suppressor of ddl that was mapped on chromosome 3 of Arabidopsis. Another component identified was MODIFIER OF DDL (MDL), a natural variation between Col and WS-2 ecotype of Arabidopsis. The variation has been mapped to an interval consisting of 37 genes on chromosome 5. MDLCol is the dominant allele and imparts a weak phenotype to ddl knockouts, whereas the recessive MDLWS-2 does not modify a strong ddl knockout phenotype. ddl MDLCol does not display abnormality in microRNA accumulation unlike ddl MDLWS-2 indicating that MDL has a function related to microRNA biogenesis.

Barley leaf rust (<I>Puccinia hordei</I> G. Otth) has been the cause of numerous and often devastating disease epidemics since the beginning of agriculture. Leaf rust is one of the most important diseases that affect barley (<I>Hordeum vulgare</I> L.) throughout the world. The pathogen <I>Puccinia hordei</I> is an obligate parasite. Symptoms of barley leaf rust may range from small chlorotic flecks to large pustules containing spores. Leaf rust epidemics reduce yields and grain quality. Deployment of resistant cultivars is one of the most effective and economical means of controlling barley leaf rust. Identification and incorporation of new and effective sources of resistance are crucial to the success of barley breeding programs. Two types of resistance have been identified. They are race-specific resistance and partial resistance. A hypersensitive reaction by the host to infection of <I>P. hordei</I> isolates lacking corresponding virulence genes is indicative of race-specific resistance that is controlled by major genes. Sixteen race-specific genes (R<I>ph</I>1 to R<I>ph</I>16) have been identified. Partial resistance is generally polygenic and is often more durable that race-specific resistance. The purpose of this research is to determine the inheritance of resistance to leaf rust in the barley experimental line VA 92-42-46, to identify the gene(s) conferring resistance, identify putative resistance related markers, and to map the gene(s) to one or more barley chromosomes using molecular markers. The Virginia barley line 92-42-46 was selected for this research project because it possesses resistance to <I>P. hordei</I> race 30, which has overcome resistance conferred by R<I>ph</I>7. Crosses were made between VA 92-42-46 and Moore, a susceptible cultivar to leaf rust. Inheritance studies were performed by screening F₂ progeny and F_2:3 families against race 8 and race 30 to determine the number of leaf rust resistance genes in VA 92-42-46. Allelism tests were performed to determine gene identity. A single dominant gene at the R<I>ph</I>5 locus or a tightly linked gene confers the resistance to P. hordei in VA 92-42-46. Two populations, 'Moore' X VA 92-42-46 and 'Bowman' X 'Magnif', were used in this study for mapping molecular markers to provide comparison and confirmation of results. 'Magnif' possesses the resistance gene R<I>ph</I>5. Bulked segregant analysis was used to identify polymorphic RFLP and SSR markers that were used for mapping in each population. Linkage analysis revealed that the R<I>ph</I>5 gene maps to barley chromosome 3 (3H) above the centromeric region in the 'Moore' X VA 92-42-46 population. These findings agree with previous research that identified linkage between R<I>ph</I>5 and R<I>ph</I>7 on chromosome 3. The results obtained in this study do not support previous research that had reported the resistance gene R<I>ph</I>5 was located on barley chromosome 7 (5H). Further research should be conducted to verify the results of this study using the 'Bowman' X 'Magnif' population. The markers screened in the region above the centromere region of barley chromosome 3 were monomorphic for the 'Bowman' X 'Magnif' population except for the marker MWG561. Therefore, additional markers above the centromere of barley chromosome 3 should be screened. / Master of Science

Heat stress adversely affects wheat production in many regions of the world and is particularly detrimental during reproductive development. The objective of this study was to identify quantitative trait loci (QTL) associated with improved heat tolerance in hexaploid bread wheat (Triticum aestivum). To accomplish this objective, an analysis of both the phenotypic and genetic responses of two recombinant inbred line (RIL) populations was conducted. RIL populations Halberd x Cutter and Halberd x Karl 92 (H/K) both derive heat tolerance from Halberd and segregate in their response to heat stress. A heat susceptibility index (HSI) was calculated from the reduction of three yield components; kernel number, kernel weight, and single kernel weight, following a three-day 38 degrees C heat stress treatment during early grain-filling. The HSI, as well as temperature depression of the main spike and flag leaf were used as measurements of heat tolerance. Genetic linkage maps were constructed for both populations and were used in combination with phenotypic data and statistical software to detect QTL for heat tolerance. In a comparison across the two across populations, seven common QTL regions were identified for HSI, located on chromosomes 1B, 3B, 4A, 5A, 5B, and 6D. Subsequent analysis of temperature depression in the H/K population identified seven QTL that co-localized for both cooler organ temperature and improved HSI. Four of the beneficial alleles at these loci were contributed Halberd. The genetic effect of combining QTL, including QHkw.tam-1B, QHkwm.tam-5A.1, and QHskm.tam-6D showed the potential benefit of selection for multiple heat tolerant alleles simultaneously. Analysis of the H/K population in the field under abiotic stress detected QTL on chromosome 3B and 5A, which were in agreement with results from the greenhouse study. The locus QYld.tam-3B was pleiotropic for both temperature depression and HSI in both experiments and was associated with higher biomass and yield under field conditions. The results presented here represent a comprehensive analysis of both the phenotypic response of wheat to high temperature stress and the genetic loci associated with improved heat tolerance and will be valuable for future understanding and improvement of heat stress tolerance in wheat.

Os objetivos do presente trabalho foram estudar a herança da resistência à antracnose foliar em milho, estimar os parâmetros genéticos e identificar marcadores moleculares ligados a genes de resistência a esta doença. Parâmetros genéticos foram estimados com base na análise de modelos de herança mista de seis gerações de quatro cruzamentos entre duas linhagens resistentes (DAS4 e DAS3) e duas linhagens suscetíveis (DAS6 e DAS22). O delineamento experimental foi o de blocos casualizados com parcelas subdivididas, com três repetições, sendo as parcelas constituídas pelos cruzamentos e as subparcelas, pelas gerações. As plantas foram inoculadas artificialmente e avaliadas em dois experimentos através de uma escala de notas de 1 a 6. Os testes de hipóteses para selecionar o modelo de herança genética e as estimativas dos parâmetros foram realizados pelo método da máxima verossimilhança. Os resultados da análise de modelos mistos indicaram que a resistência é controlada por um gene de efeito maior em todos os cruzamentos e experimentos avaliados e também por poligenes, em pelo menos um dos experimentos. A ação gênica é aditiva e dominante, com predominância de efeitos genéticos aditivos. O mapeamento de QRLs foi realizado utilizando 141 indivíduos F1RC1 do cruzamento (DAS6 x DAS4) x DAS6, com base na avaliação fenotípica das suas famílias, em dois experimentos. O delineamento experimental foi o látice 12 x 12, incluindo as famílias, genitores e híbrido, com 3 repetições. As plantas foram inoculadas artificialmente e avaliadas através de uma escala de notas de 1 a 6. O mapeamento de QRLs, utilizando marcadores microssatélites e AFLPs e análise de bulks segregantes para detecção de marcadores candidatos, foi realizado pela análise de regressão linear múltipla (RLM) e pelo mapeamento por intervalo composto (MIC). Ambas metodologias de análise identificaram pelo menos um QRL no cromossomo 10 em cada experimento. Também foram detectados QRLs nos cromossomos 2, 3 e 5, apenas pela RLM. Os QRLs identificados pela RLM explicaram 25,7%, 23,3% e 24,5% da variação fenotípica no experimento 1, no experimento 2 e na análise conjunta, respectivamente. Já os QRLs identificados pelo MIC explicaram 28,9%, 32,3% e 31,0% da variação fenotípica no experimento 1, no experimento 2 e na análise conjunta, respectivamente. A análise de bulks segregantes permitiu a detecção apenas dos QRLs de efeitos fenotípicos mais expressivos localizados no cromossomo 10. Na maioria dos QRLs detectados, os alelos de resistência provieram do genitor resistente. A identificação destes QRLs oferece uma significativa contribuição para o entendimento da resistência de milho à antracnose foliar, podendo levar à identificação de genes e elucidação dos mecanismos envolvidos na expressão da resistência. / The objectives of this work were to study the inheritance of resistance to anthracnose leaf blight, estimate the genetic parameters, and identify molecular markers associated with resistance genes to this disease. Genetic parameters were estimated based on the analysis of mixed inheritance models in six generations of four crosses between two resistant (DAS4 and DAS3) and two susceptible inbred lines (DAS6 and DAS22). The experimental design consisted of randomized blocks containing split-plots, with three replicates, where the plots were represented by crosses and the subplots were the generations. The plants were inoculated artificially and evaluated in two experiments by means of a rating scale from 1 to 6. The hypothesis testing to select the genetic inheritance model and parameter estimates were obtained by the maximum likelihood method. The results from the mixed models analysis indicated that resistance is controlled by a major gene in all crosses and experiments evaluated, and also by polygenes in at least one experiment. The genetic action is additive and dominant, with predominance of additive genetic effects. QRL mapping was performed using 141 F1RC1 individuals from the (DAS6 × DAS4) × DAS6 cross, based on the phenotypic evaluation of their families, in two experiments. The experimental design was a 12 × 12 lattice which included the families, parents, and the hybrid, with 3 replicates. The plants were inoculated artificially and evaluated by means of a rating scale from 1 to 6. QRL mapping, using microsatellites and AFLPs markers, and bulked segregant analysis to detect candidate markers was performed by multiple linear regression analysis (MLR) and by composite interval mapping (CIM). Both methodologies of analysis identified at least one QRL in chromosome 10 in each experiment. QRLs were also detected, by MLR only, in chromosomes 2, 3 and 5. The QRLs identified by MLR explained 25.7%, 23.3%, and 24.5% of the phenotypic variation in the experiment 1, in the experiment 2 and in the joint analysis, respectively. The QRLs identified by CIM, however, explained 28.9%, 32.3%, and 31.0% of the phenotypic variation in the experiment 1, in the experiment 2 and in the joint analysis, respectively. The bulked segregant analysis only allowed the detection of QRLs that showed the more expressive phenotypic effects located in chromosome 10. In most detected QRLs, the resistance alleles came from the resistant parent. The identification of these QRLs offers a significant contribution to an understanding of resistance to anthracnose leaf blight in maize, and could lead to the identification of genes and to an elucidation of the mechanisms involved in the expression of resistance.

Os objetivos do presente trabalho foram estudar a herança da resistência à antracnose foliar em milho, estimar os parâmetros genéticos e identificar marcadores moleculares ligados a genes de resistência a esta doença. Parâmetros genéticos foram estimados com base na análise de modelos de herança mista de seis gerações de quatro cruzamentos entre duas linhagens resistentes (DAS4 e DAS3) e duas linhagens suscetíveis (DAS6 e DAS22). O delineamento experimental foi o de blocos casualizados com parcelas subdivididas, com três repetições, sendo as parcelas constituídas pelos cruzamentos e as subparcelas, pelas gerações. As plantas foram inoculadas artificialmente e avaliadas em dois experimentos através de uma escala de notas de 1 a 6. Os testes de hipóteses para selecionar o modelo de herança genética e as estimativas dos parâmetros foram realizados pelo método da máxima verossimilhança. Os resultados da análise de modelos mistos indicaram que a resistência é controlada por um gene de efeito maior em todos os cruzamentos e experimentos avaliados e também por poligenes, em pelo menos um dos experimentos. A ação gênica é aditiva e dominante, com predominância de efeitos genéticos aditivos. O mapeamento de QRLs foi realizado utilizando 141 indivíduos F1RC1 do cruzamento (DAS6 x DAS4) x DAS6, com base na avaliação fenotípica das suas famílias, em dois experimentos. O delineamento experimental foi o látice 12 x 12, incluindo as famílias, genitores e híbrido, com 3 repetições. As plantas foram inoculadas artificialmente e avaliadas através de uma escala de notas de 1 a 6. O mapeamento de QRLs, utilizando marcadores microssatélites e AFLPs e análise de bulks segregantes para detecção de marcadores candidatos, foi realizado pela análise de regressão linear múltipla (RLM) e pelo mapeamento por intervalo composto (MIC). Ambas metodologias de análise identificaram pelo menos um QRL no cromossomo 10 em cada experimento. Também foram detectados QRLs nos cromossomos 2, 3 e 5, apenas pela RLM. Os QRLs identificados pela RLM explicaram 25,7%, 23,3% e 24,5% da variação fenotípica no experimento 1, no experimento 2 e na análise conjunta, respectivamente. Já os QRLs identificados pelo MIC explicaram 28,9%, 32,3% e 31,0% da variação fenotípica no experimento 1, no experimento 2 e na análise conjunta, respectivamente. A análise de bulks segregantes permitiu a detecção apenas dos QRLs de efeitos fenotípicos mais expressivos localizados no cromossomo 10. Na maioria dos QRLs detectados, os alelos de resistência provieram do genitor resistente. A identificação destes QRLs oferece uma significativa contribuição para o entendimento da resistência de milho à antracnose foliar, podendo levar à identificação de genes e elucidação dos mecanismos envolvidos na expressão da resistência. / The objectives of this work were to study the inheritance of resistance to anthracnose leaf blight, estimate the genetic parameters, and identify molecular markers associated with resistance genes to this disease. Genetic parameters were estimated based on the analysis of mixed inheritance models in six generations of four crosses between two resistant (DAS4 and DAS3) and two susceptible inbred lines (DAS6 and DAS22). The experimental design consisted of randomized blocks containing split-plots, with three replicates, where the plots were represented by crosses and the subplots were the generations. The plants were inoculated artificially and evaluated in two experiments by means of a rating scale from 1 to 6. The hypothesis testing to select the genetic inheritance model and parameter estimates were obtained by the maximum likelihood method. The results from the mixed models analysis indicated that resistance is controlled by a major gene in all crosses and experiments evaluated, and also by polygenes in at least one experiment. The genetic action is additive and dominant, with predominance of additive genetic effects. QRL mapping was performed using 141 F1RC1 individuals from the (DAS6 × DAS4) × DAS6 cross, based on the phenotypic evaluation of their families, in two experiments. The experimental design was a 12 × 12 lattice which included the families, parents, and the hybrid, with 3 replicates. The plants were inoculated artificially and evaluated by means of a rating scale from 1 to 6. QRL mapping, using microsatellites and AFLPs markers, and bulked segregant analysis to detect candidate markers was performed by multiple linear regression analysis (MLR) and by composite interval mapping (CIM). Both methodologies of analysis identified at least one QRL in chromosome 10 in each experiment. QRLs were also detected, by MLR only, in chromosomes 2, 3 and 5. The QRLs identified by MLR explained 25.7%, 23.3%, and 24.5% of the phenotypic variation in the experiment 1, in the experiment 2 and in the joint analysis, respectively. The QRLs identified by CIM, however, explained 28.9%, 32.3%, and 31.0% of the phenotypic variation in the experiment 1, in the experiment 2 and in the joint analysis, respectively. The bulked segregant analysis only allowed the detection of QRLs that showed the more expressive phenotypic effects located in chromosome 10. In most detected QRLs, the resistance alleles came from the resistant parent. The identification of these QRLs offers a significant contribution to an understanding of resistance to anthracnose leaf blight in maize, and could lead to the identification of genes and to an elucidation of the mechanisms involved in the expression of resistance.

Doctor of Philosophy / Department of Agronomy / Gina Brown-Guedira / Allan K. Fritz / Aegilops tauschii, the D-genome donor of modern wheat, has served as an important source of genetic variation in wheat breeding. The objective of this research was to characterize and utilize multiple Ae. tauschii-derived pest resistance genes contained in the germplasm KS96WGRC40. Two Ae. tauschii-derived genes, H23 and Cmc4, provide resistance to the Hessian fly (HF) and wheat curl mite (WCM), respectively. A linkage analysis of a testcross population estimated 32.67% recombination between H23 and Cmc4 on chromosome 6DS in wheat indicating that the two genes are not tightly linked as previous mapping reports show. Haplotype data of recombinant lines and physical mapping of linked microsatellite markers located Cmc4 distal to H23. Haplotype data indicated that both KS89WGRC04 and KS96WGRC40 have the distal portion of 6DS derived from Ae. tauschii. Microsatellite primer pairs BARC183 and GDM036 were more useful than the previously published linked markers in identifying lines carrying Cmc4 and H23, respectively. Through phenotypic selection and advancement within the testcross population, three TC1F2:4 lines were identified as homozygous resistant for H23 and Cmc4 and have the complete terminal segment of 6DS from Ae. tauschii. Two lines are more desirable than the original germplasm releases and can serve as a source of resistance to both HF and WCM in an elite background. A linkage analysis of a segregating recombinant inbred line population identified an Ae. tauschii-derived gene of major effect conferring resistance to Septoria leaf blotch (STB) and another Ae. tauschii-derived gene of major effect conferring resistance to soil-borne wheat mosaic virus (SBWMV) in the germplasm KS96WGRC40. The STB resistance gene in KS96WGRC40 is located in the distal 40% of the short arm of chromosome 7D flanked by microsatellite markers Xgwm044 and Xbarc126. Two previously reported STB genes, Stb4 and Stb5, have been mapped to 7DS in the same region as the STB resistance gene in KS96WGRC40. The uniqueness of the STB resistance genes on 7DS is questionable. The SBWMV resistance gene in KS96WGRC40 is located on chromosome 5DL linked to microsatellite marker Xcfd010. The SBWMV resistance gene within KS96WGRC40 was derived from TA2397 via KS95WGRC33.

No description available.