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Title
A nitrogen-responsive small peptide signaling mechanism modulates plant root system architecture
Creator
Lay-Pruitt, Katerina Sibala
Date
2021
Collection
Electronic Theses & Dissertations
Description
The plant root system changes dynamically in response to environmental cues. Plants utilize their root system for uptake of essential mineral nutrients that are heterogeneously distributed in the soil environment. Nutrient-dependent modulation of root system architecture (RSA) traits such as primary root growth, lateral root emergence, and the angles at which these roots grow allows for optimization of nutrient acquisition. Among signaling pathways by which plants may sense the availability...
Show moreThe plant root system changes dynamically in response to environmental cues. Plants utilize their root system for uptake of essential mineral nutrients that are heterogeneously distributed in the soil environment. Nutrient-dependent modulation of root system architecture (RSA) traits such as primary root growth, lateral root emergence, and the angles at which these roots grow allows for optimization of nutrient acquisition. Among signaling pathways by which plants may sense the availability of nutrients from the environment, small signaling peptide (SSP) pathways play important roles in optimizing root functions. These SSP pathways may further regulate molecular processes underlying RSA, such as the biosynthesis and transport of the major plant growth hormone, auxin. Characterization of these nutrient-responsive SSP pathways is thus of great importance and critical for understanding plant development in nutrient-poor environments. For my dissertation, I have identified and characterized a nitrogen (N)-responsive SSP pathway modulating root gravitropic response and lateral root development. Co-regulation of these RSA components by this module is proposed to prevent root outgrowth into N-poor regions and drive deeper root growth towards mobile nitrate (NO3-) resources stratified deeper in the soil profile. First, I show that a signaling pathway involving the CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) family of peptides and the CLAVATA1 (CLV1) receptor kinase, which is involved in N-dependent repression of lateral root emergence, also enhances root gravitropic response under N-limited conditions. Transcriptomic profiling of a clv1 mutant and CLE3 overexpressing lines identified Arabidopsis thaliana CENTRORADIALIS (ATC), a mobile protein previously characterized for its role in flowering regulation, as a downstream target of CLE-CLV1 signaling. Loss of ATC function significantly weakens root gravitropic response and has a moderate impact on lateral root emergence under low NO3- availability. ATC promoter activity and protein localization are also detected throughout the phloem and in the root columella cells, which are major centers for gravity sensing. Second, I demonstrate the relevance of ATC function on the molecular processes underlying root gravitropic response. While mutation in ATC does not impact gravity sensing via amyloplast sedimentation, it does inhibit the asymmetric transport of auxin needed for gravitropic bending. I determine that this occurs via the significant reduction of the PIN3 auxin efflux transporter in the vasculature and root tip of atc mutant lines. Lastly, I examine how the known roles of ATC in floral development could be implicated in root developmental processes. ATC binds to phosphatidic acid and phosphatidylserine, which is contrary to the binding capacity of its homolog FLOWERING LOCUS T (FT) to phosphatidylcholine and may contribute to its activity in N-limited environments. I also investigate the interaction of ATC and the transcription factor FD in the transcriptional regulation of PIN3. Although FD appears to have an impact on root gravitropic response, FD inhibits the expression of PIN3, suggesting potentially complex control of this gene via floral regulatory components. Taken together, the results presented in this dissertation contribute greatly to our understanding of how plant root architecture alters in response to N. These results can be further utilized in plant engineering strategies to regulate root growth in nutrient-limited soils.
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Title
USING FRAGARIA AS A MODEL SYSTEM FOR THE STUDY OF SUBGENOME DOMINANCE AND ADAPTATION IN CROPS
Creator
Alger, Elizabeth
Date
2021
Collection
Electronic Theses & Dissertations
Description
Polyploidy, or the presence of three or more complete genomes in a single organism, has occurred frequently in plants, especially in the angiosperm lineage. Allopolyploids, or polyploids resulting from the merging of different genomes in an interspecific hybrid, have often been shown to experience subgenome dominance. Subgenome dominance is the phenomenon where there is bias in the gene loss and expression between the different genomes in a polyploid, known as subgenomes. Despite the...
Show morePolyploidy, or the presence of three or more complete genomes in a single organism, has occurred frequently in plants, especially in the angiosperm lineage. Allopolyploids, or polyploids resulting from the merging of different genomes in an interspecific hybrid, have often been shown to experience subgenome dominance. Subgenome dominance is the phenomenon where there is bias in the gene loss and expression between the different genomes in a polyploid, known as subgenomes. Despite the prevalence of polyploids and subgenome dominance, little is known about the factors and mechanisms that influence this process. Strawberry (Fragaria sp.) is emerging as a powerful model system to investigate polyploid subgenome dominance evolution due to the recent identification of the four extant diploid progenitor species of the cultivated octoploid strawberry (Fragaria x ananassa). Having the diploid progenitors in hand allows us to identify differences between the dominant subgenome, F. vesca, and the other three progenitors that may have an impact of subgenome dominance. One possible factor is transposable element (TE) abundance, as low TE density has been consistently associated with the dominant subgenome in allopolyploids. Epigenetic silencing of TEs by DNA methylation to suppress TE activity has been shown to result in decreased expression of neighboring genes and this lowered gene expression may affect the establishment of subgenome dominance. F. vesca will be used as a diploid model for the study of subgenome dominance in strawberry where I can examine how TE abundance and other factors influence gene expression in a single accession and in hybrid crosses between different accessions. Tracking changes in gene expression in the hybrids will allow us to examine how genomes with difference sizes and genomic factors interact. The results and insights observed from this study can then be applied to subgenome dominance research in octoploid strawberry. In addition to the germplasm and genomic resources, strawberries are also a high value crop and the loss of their production due to (a)biotic stressors results in the loss of millions of United States dollars annually. Using a population of octoploid strawberries segregating for salt tolerance, I will identify candidate genes related to salt tolerance. Together this work will identify factors and mechanisms related to subgenome dominance and use genotypic data in a practical breeding context.
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Title
DECIPHERING THE GENETIC BASIS OF SOLANUM CHACOENSE MEDIATED COLORADO POTATO BEETLE (LEPTINOTARSA DECEMLINEATA) RESISTANCE AND SELF- FERTILITY IN A DIPLOID SOLANUM CHACOENSE RECOMBINANT INBRED LINE POPULATION
Creator
Kaiser, Natalie
Date
2021
Collection
Electronic Theses & Dissertations
Description
The Colorado potato beetle (Leptinotarsa decemlineata) is the most widespread and destructive insect defoliator pest of potato and its control has historically been achieved through the use of insecticide. The diploid potato species Solanum chacoense has been utilized for over four decades in an attempt to introgress glycoalkaloid-based insect resistance into cultivated tetraploid potato. Despite these efforts, insect resistant cultivars have not been achieved, due in part to the complex...
Show moreThe Colorado potato beetle (Leptinotarsa decemlineata) is the most widespread and destructive insect defoliator pest of potato and its control has historically been achieved through the use of insecticide. The diploid potato species Solanum chacoense has been utilized for over four decades in an attempt to introgress glycoalkaloid-based insect resistance into cultivated tetraploid potato. Despite these efforts, insect resistant cultivars have not been achieved, due in part to the complex genetics underlying the trait. The creation of inbred diploid lines would allow more efficient examination and deployment of this economically important trait. We introduced self-compatibility into diploid insect resistant S. chacoense germplasm and developed the first potato recombinant inbred line (RIL) population to study, understand and deploy this mechanism of host-plant insect resistance in cultivated, diploid breeding lines.We first examined the genetic features underlying leptine glycoalkaloid mediated Colorado potato beetle host plant resistance in the F2 generation derived from a cross between S. chacoense lines USDA8380-1 (80-1) and M6. Using biparental linkage mapping, a major overlapping QTL region with dominant effects was identified on chromosome 2 explaining 49.3% and 34.1% of the variance in Colorado potato beetle field resistance and leptine accumulation, respectively. Bulk segregant whole genome sequencing of the same F2 population detected QTL associated with Colorado potato beetle resistance on chromosomes 2, 4, 6, 7, and 12. Candidate genes within these QTL regions were identified by weighted gene co-expression network analysis of parental lines and resistant and susceptible F2 individuals. Second, we exploited M6-mediated self-compatibility and established vigorous, F5 inbred diploid lines to further examine loci associated with Colorado potato beetle resistance and explore the practicality of inbreeding in diploid potato. F5 inbred lines carrying Colorado potato beetle resistance equivalent to the resistant donor parent were created without field selection during the inbreeding process. We report that the ratio of acetylated to non-acetylated glycoalkaloids measured under greenhouse conditions is a powerful metabolite marker to predict field performance without incurring the costs of conducting a Colorado potato beetle field trial. Leptine production was successfully introduced into diploid breeding germplasm. Single nucleotide polymorphism (SNP) genotyping coupled with stylar analysis of pollen tube growth and self-fertility phenotyping of the F4 and F5 generations revealed that multiple factors mediate the self-compatible response in this RIL population. Third, we assessed the initial transcriptional and metabolite response to Colorado potato beetle herbivory in beetle resistant and beetle susceptible S. chacoense lines over a 48-hour time course. To facilitate genome editing modification of the leptine biosynthesis pathway, we characterized the allelic variation between S. chacoense 80-1 and M6 in a candidate leptine biosynthesis gene identified by transcriptional profiling. This work highlights the challenges of establishing inbred germplasm, reinforces the complexity of selecting for self-fertility in diploid potato, and lays the foundation for optimization of potato RIL development. The availability of highly homozygous Colorado potato beetle resistant lines will enable further genomic inquiry of the loci contributing to this trait and will facilitate rapid deployment of beetle resistant diploid potato varieties.
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Title
A FRAMEWORK FOR BIOLOGICAL DATA INTEGRATION AND FEATURE SELECTION IN LARGE DATA SETS
Creator
Gonzalez-Reymundez, Agustin
Date
2021
Collection
Electronic Theses & Dissertations
Description
The increasing volume of high-dimensional biological data (omics) has intensified the discovery of thousands of biomarkers across the different fundamental components of the cell (e.g., genome, transcriptome, proteome, epigenome) and allowed the characterization of complex phenotypes (e.g., metabolome, imaginome, phenome). However, the ability to integrate omics into informative results is constantly challenged by a seemingly ever-increasing volume of data. Furthermore, huge data sizes impose...
Show moreThe increasing volume of high-dimensional biological data (omics) has intensified the discovery of thousands of biomarkers across the different fundamental components of the cell (e.g., genome, transcriptome, proteome, epigenome) and allowed the characterization of complex phenotypes (e.g., metabolome, imaginome, phenome). However, the ability to integrate omics into informative results is constantly challenged by a seemingly ever-increasing volume of data. Furthermore, huge data sizes impose a tradeoff between how complex an omic integration algorithm can be and how much data it can handle (e.g., how fast can the algorithm be scaled to integrate large data sizes). In this dissertation, we explore statistical frameworks to face the challenges of modern omic data, including the integration of high-dimensional data of large sample sizes. We have developed a novel framework of competitive analytical performance compared with existing methods but suitable for omic data reaching biobank scales (i.e., hundreds of thousands of samples and variables). We implemented this method as an R package and showed its application on two traits of a complex molecular basis: cancer and regulation of energy intake and expenditure. In chapter one, we review the technologies and methods used to generate and integrate omic data. Chapter two describes our novel method and software of omic integration, shows examples in synthetic data, and evaluates its computational and analytical performance. Chapter three presents an application of our method to reveal a novel pan-cancer classification of tumors beyond the tissue of origin, regulated by distinct sets of molecular signatures. In chapter four, we present an application of our method to integrate phenomics data and identify patterns of energy balance regulated by genomic variation. Finally, in chapter five, we offer general conclusions to the entire thesis.
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Title
Genomic basis of electric signal variation in African weakly electric fish
Creator
Losilla-Lacayo, Mauricio
Date
2021
Collection
Electronic Theses & Dissertations
Description
A repeated theme in speciation is reproductive isolation centered around divergence in few, highly variable traits, specially in cases without strong geographic isolation and high speciation rates. Understanding the genomic basis of highly variable traits that are key to speciation is a major goal of evolutionary biology, because they can characterize crucial drivers and foundations of the speciation process. African weakly electric fish (Mormyridae) are a decidedly speciose clade of teleost...
Show moreA repeated theme in speciation is reproductive isolation centered around divergence in few, highly variable traits, specially in cases without strong geographic isolation and high speciation rates. Understanding the genomic basis of highly variable traits that are key to speciation is a major goal of evolutionary biology, because they can characterize crucial drivers and foundations of the speciation process. African weakly electric fish (Mormyridae) are a decidedly speciose clade of teleost fish, and their electric organ discharges (EODs) are highly variable traits central to species divergence. However, little is known about the genes and celullar processes that underscore EOD variation. In this dissertation, I employ RNAseq and Nanopore sequencing to study the genomic basis of electric signal variation in mormyrids. In Chapter 1, I take a transcriptome-wide approach to describe the molecular basis of electric signal diversity in species of the mormyrid genus Paramormyrops, divergent for EOD complexity, duration and polarity. My results emphasize genes that influence the shape and structure of the electrocyte cytoskeleton, membrane, and extracellular matrix, and the membrane’s physiological properties. In Chapter 2, I compare gene expression patterns between electric organs that produce long vs short EODs. The results strongly support known aspects of morphological and physiological bases of EOD duration, and for the first time I identified specific genes and broad cellular processes expected to that alter morphological and physiological properties of electrocytes, most striking among these is the differential expression of multiple potassium voltage-gated channels. These two chapters independently identified the gene epdl2 as of interest for EOD divergence. In Chapter 3, I study the molecular evolutionary history of epdl2 in Mormyridae, with emphasis on Paramormyrops. My results suggest that three rounds of gene duplication produced four epdl2 paralogs in a Paramormyrops ancestor. In addition, I identify ten sites in epdl2 expected to have experienced strong positive selection in paralogs and implicate them in key functional domains. Overall, the results of this dissertation greatly solidify and expand our understanding of how the genome underpins changes to electrocytes, and in turn, divergence in their electric signals, a highly variable trait that may facilitate speciation in African weakly electric fish. This work provides an evidence-grounded list of candidate genes for functional analyses aimed to corroborate their contribution to the EOD phenotype.
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Title
Developing Reverse Genetic Tools in Weakly Electric Fish : Investigating Electric Organ in vivo scn4aa Function Through CRISPR Knockouts and Morpholino Knockdowns
Creator
Constantinou, Savvas James
Date
2021
Collection
Electronic Theses & Dissertations
Description
The ability to determine gene function allows research to progress at one of the finest scales in biology and is a goal in electric fish research. Reverse genetics allows researchers to determine gene function and would aid the electric fish community in beginning to answer some of the broadest and most complicated questions in biology such as linking genotype to phenotype and understanding the processes that lead to biological diversity. In this dissertation, I describe the development of...
Show moreThe ability to determine gene function allows research to progress at one of the finest scales in biology and is a goal in electric fish research. Reverse genetics allows researchers to determine gene function and would aid the electric fish community in beginning to answer some of the broadest and most complicated questions in biology such as linking genotype to phenotype and understanding the processes that lead to biological diversity. In this dissertation, I describe the development of two major reverse genetic tools for use in the electric fish system: CRISPR/Cas9 genome editing and morpholinos. To develop these tools, I also produced protocols for in-vitro breeding, husbandry, and single-cell embryo microinjections. In the first chapter, I describe in-vitro breeding, husbandry, and single-cell embryo microinjections and demonstrate that CRISPR/Cas9 is a promising tool for future electric fish research by targeting nonsense mutations to scn4aa in the mormyrid Brienomyrus brachyistius and gymnotiform Brachyhypopomus gauderio, two independently evolved lineages of weakly electric fish, resulting in a reduction in the electric organ discharge amplitude. In the second chapter, I provide electric fish researchers with a detailed analysis of our many successes and failures applying CRISPR/Cas9 methods to this system and discuss future suggestions on how to best apply them to novel electric fish research. In the third chapter, I describe my efforts to utilize vivo-morpholinos in mormyrid electric fish. While a single early pilot study I performed demonstrated vivo-morpholinos can reduce target gene mRNA levels and cause a phenotypic effect, my efforts to replicate these findings demonstrate inconsistent performance: control vivo-morpholino and scn4aa targeting vivo-morpholino injected fish had indistinguishable effects on electric organ discharge amplitude. Due to additional concerns of toxicity, I suggest morpholinos are not an ideal reverse genetic tool in Brienomyrus brachyistius and should only be utilized for future research with caution.
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Title
Genetic And Genetic By Environment Effects On Tar Spot Resistance And Hybrid Yield In Maize
Creator
Trygestad, Blake
Date
2021
Collection
Electronic Theses & Dissertations
Description
The phenotype of any plant can be broken down into the three primary sources of variation, genetic (G), environment (E), and genetic by environmental interaction (GxE). Producers and researchers alike will harness repeatable G and GxE effects to maximize their resource efficiency. This study studied the G and GxE effects in the biotic stress of the fungi Phyllachora maydis and the environment patterns in advanced yield trial data. In rating 800 genotypes over two seasons, we genetically...
Show moreThe phenotype of any plant can be broken down into the three primary sources of variation, genetic (G), environment (E), and genetic by environmental interaction (GxE). Producers and researchers alike will harness repeatable G and GxE effects to maximize their resource efficiency. This study studied the G and GxE effects in the biotic stress of the fungi Phyllachora maydis and the environment patterns in advanced yield trial data. In rating 800 genotypes over two seasons, we genetically mapped and identified over 100 significant Single Nucleotide Polymorphisms (SNPs) associated with tar spot resistance using a genome-wide association study. We then conducted genomic prediction, which was 81.5% accurate for predicting tar spot severity within the location and 48% accurate in predicting disease resistance in a new environment. Also, using Genetic and Genotype x Environment (GGE) biplots, we investigated environmental patterns of nine locations in three maturity Zones in the advanced yield trials in the Michigan Yield Performance Trials. First, we identified two locations, one in the late and one in the mid maturity zone, with equal G and GxE effects and should be removed. Then, using a sliding window of year combinations, we analyzed the optimal number of replications needed across the three maturity zones. We determined that an average of three replications are needed to achieve 75% of the maximum repeatability across the zones.
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Title
INTEGRATED ANALYSIS OF GENETIC MARKER, MIRNA, AND MRNA DATA TO UNRAVEL MECHANISMS CONTROLLING GROWTH AND MEAT QUALITY TRAITS IN PIGS
Creator
Daza, Kaitlyn R.
Date
2021
Collection
Electronic Theses & Dissertations
Description
Determining mechanisms regulating complex traits in pigs is essential to improve the production efficiency of this globally important protein source. MicroRNAs (miRNAs) are a class of non-coding RNAs known to post-transcriptionally regulate gene expression affecting numerous tissues and phenotypes, including those important to the pig industry. However, further research is needed to characterize the miRNAs expressed in pig skeletal muscle and assess their impact on the regulation of growth,...
Show moreDetermining mechanisms regulating complex traits in pigs is essential to improve the production efficiency of this globally important protein source. MicroRNAs (miRNAs) are a class of non-coding RNAs known to post-transcriptionally regulate gene expression affecting numerous tissues and phenotypes, including those important to the pig industry. However, further research is needed to characterize the miRNAs expressed in pig skeletal muscle and assess their impact on the regulation of growth, carcass composition, and meat quality traits. Additionally, little is known about the genetic architecture controlling miRNA expression in pigs, which can be elucidated by combining high-density genotypic data with miRNA expression profiles from the same animals in the mapping of miRNA expression quantitative trait loci (miR-eQTL). This analysis reveals associations between genomic regions harboring single nucleotide polymorphisms (SNPs) and variation in the expression of miRNAs. By integrating mRNA expression profiles and phenotypic data from the same individuals, putative regulatory relationships can be revealed underlying variation in phenotypes relevant to pig production. In this study, our objectives were to profile and characterize the small RNA population present in longissimus dorsi (LD) skeletal muscle samples from a F2 Duroc x Pietrain resource population, and to conduct an integrated miR-eQTL analysis to identify regulators of miRNA expression and candidate genes regulating phenotypic traits in adult pig skeletal muscle.MicroRNA expression profiling was performed on total RNA extracted from (LD) muscle samples from 174 F2 pigs. The composition of small RNA classes present in this dataset was characterized through a series of homology searches against human, mouse, and pig databases. MicroRNA quantification and novel miRNA prediction was conducted, profiling the abundance of 295 known mature pig miRNAs and producing 27 unique candidate novel miRNA precursors. The 295 miRNA expression profiles were subsequently used as response variables in a GBLUP-based GWA analysis. Results for associating these miRNAs with 36,292 SNPs identified 315 significant miRNA-SNP associations (FDR < 0.05), comprising 23 significant eQTL peaks associated with 17 unique miRNAs. Five of the 23 miR-eQTL peaks were defined as local-acting, meaning the genomic positions of the significantly associated SNPs comprising the miR-eQTL peak overlapped that of the miRNA precursor transcript. We then investigated the potential effects of these miRNAs through miRNA target prediction, correlation, and colocalization analyses. Notably, one miR-eQTL miRNA exhibiting a strong local¬-acting miR-eQTL, miR-874, had predicted target genes colocalizing with previously identified phenotypic QTL for 12 production traits including backfat thickness, dressing percentage, muscle pH at 24 h post-mortem, and cook yield. The results of this study revealed putative pig-novel miRNAs for further study and validation, contributing to our understanding of the miRNA landscape present in adult pig skeletal muscle. Additionally, we identified genomic regions underlying variation in miRNA expression, and candidate miRNAs and genes for future investigation of their regulatory effects on growth, carcass composition, and meat quality traits of importance to the global pig industry.
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Title
Germplasm Evaluation of Common Beans for End-Use Quality : Seed Coat Color and Cooking Time of Yellow Beans and Paste Qualities of Michigan Bean Varieties
Creator
Sadohara, Rie
Date
2021
Collection
Electronic Theses & Dissertations
Description
Consumption of dry beans is low in the U.S. and the genetic improvement of end-use quality traits of dry beans and the development of new uses for beans offer potential opportunities to increase consumption. This research aimed to 1) characterize a yellow bean diversity panel for seed coat color and cooking time, 2) investigate the G×E and inheritance of seed coat color and cooking time of the diversity panel via genome-wide association (GWA), 3) evaluate the suitability of MI-bred bean...
Show moreConsumption of dry beans is low in the U.S. and the genetic improvement of end-use quality traits of dry beans and the development of new uses for beans offer potential opportunities to increase consumption. This research aimed to 1) characterize a yellow bean diversity panel for seed coat color and cooking time, 2) investigate the G×E and inheritance of seed coat color and cooking time of the diversity panel via genome-wide association (GWA), 3) evaluate the suitability of MI-bred bean varieties for sweet paste qualities, and 4) inform bean breeders of the food industry’s needs for bean flour. A Yellow Bean Collection with 295 lines grown in MI and NE were evaluated for seed coat color, post-harvest darkening, and cooking time. Machine-learning was used to develop a procedure to automatically exclude hilum and corona areas in bean images and extract L*a*b* color values from seed coat. Color values, post-harvest darkening, hilum, and corona colors mapped to the ground factor P gene. Cooking time evaluation identified 20 lines that cooked within 20 min in MI and five lines that cooked within 31 min in higher-altitude NE. GWA identified a polygalacturonase gene as a candidate gene for cooking time. G×E was significant for seed coat color and cooking time, and MI-grown seeds were generally darker and faster-cooking than NE-grown. Six white bean varieties and one cranberry genotype were evaluated for sweet bean paste qualities: paste yield, color, stickiness, and flavor. Large-seeded white varieties performed similar or higher for many of the quality traits than Hime, an industry standard, showing their potential as a novel food application in the U.S. An electronic survey was conducted targeting U.S. food industry professionals who use pulse and/or wheat flour. Seventy-five valid responses were collected, and flavor and functionality rather than lectins or gluten contamination were the most important challenges in utilizing bean flour. These studies will be useful in selecting germplasm for favorable qualities to consumers such as bright yellow color, non-darkening after storage, fast cooking time, bean paste qualities, and bean flour attributes.
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Title
Assessment of state-specific DNA methylation patterns to improve functional annotation of farm animal genomes
Creator
Corbett, Ryan James
Date
2021
Collection
Electronic Theses & Dissertations
Description
Over the past several decades, genetic advancements in the domestic pig (Sus scrofa) and other farm animal species have resulted in increased economic output and expanded use of these organisms as biomedical models to study human disease. However, limited functional annotation of the porcine genome—particularly in non-coding regulatory regions—hinders both identification of causal genes for complex traits and translational research capabilities. The Functional Annotation of Animal Genomes...
Show moreOver the past several decades, genetic advancements in the domestic pig (Sus scrofa) and other farm animal species have resulted in increased economic output and expanded use of these organisms as biomedical models to study human disease. However, limited functional annotation of the porcine genome—particularly in non-coding regulatory regions—hinders both identification of causal genes for complex traits and translational research capabilities. The Functional Annotation of Animal Genomes consortium seeks to map functional elements in domesticated animal genomes in part by performing sequencing assays to characterize the animal epigenome, as specific chromatin modifications have been shown to be predictive of regulatory regions. DNA methylation is the most ubiquitous epigenetic modification made to the DNA molecule, and in mammals occurs almost exclusively at cytosines in CpG dinucleotides. DNA methylation exerts regulatory effects through numerous mechanisms, including the occlusion of transcription factors at activating regulatory regions, and as such has been shown to play important roles in establishing spatiotemporal gene expression. Furthermore, differential methylation has been associated with genomic imprinting and stress-induced physiological changes in mammals. Assessment of DNA methylation in the pig and other farm animal species has thus far been limited in scope. In this dissertation, I have characterized state-specific DNA methylation patterns in farm animal genomes across a diverse collection of cell types, developmental stages, and environmental conditions, to enhance understanding of epigenetic gene regulation in livestock and poultry. First, I demonstrate that sorted porcine immune cells exhibit unique DNA methylation landscapes that are strongly correlated with local and distal gene expression as well as binding sites for transcription factors regulating immune cell-specific functions. The co-localization of immune cell differentially methylated regions with GWAS SNPs for immune-related traits supports the use of epigenomics assays to increase functional annotation of economically relevant genomic regions. Second, I show that development of four porcine fetal tissues (whole brain, liver, loin muscle, and placenta) is associated with increased differentiation of DNA methylation profiles that likely contributes to tissue-specific transcriptomes and transcription factor regulatory potential. I also report widespread allele-biased methylation in fetal tissues associated with breed-specific gene regulation as well as putative regions of genomic imprinting events. Third, I characterize associations between environmental stimuli and DNA methylation patterns in two studies. I show that piglet weaning correlates with changes in peripheral blood mononuclear cell DNA methylation, and that increased weaning stress is associated with increased methylation and decreased expression of T cell-enriched genes, suggesting a diminished adaptive immune response. Lastly, I assess the impact of broiler chick incubation parameters on cardiac DNA methylation and observe significant temperature-associated differential methylation of genes involved in heart morphogenesis. I identified differentially methylated and expressed genes between temperature treatments that may influence environment-driven differences in cardiovascular development. In conclusion, I have performed the most expansive survey of whole-genome DNA methylation in farm animal species to date and have identified thousands of putative regulatory elements influencing state-specific gene and phenotype expression. These data will be a valuable resource for future functional annotation efforts seeking to identify mechanistic links between genetic and phenotypic variation in animal species.
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Title
EVALUATING GENOMIC ESTIMATES AND RECONSTRUCTED PEDIGREES AS ASSESSMENT TECHNIQUES FOR SEA LAMPREY POPULATIONS
Creator
Weise, Ellen M.
Date
2021
Collection
Electronic Theses & Dissertations
Description
Sea lamprey (Petromyzon marinus) are a harmful invasive species in the Great Lakes. Their invasion resulted in the decimation of native fish populations. A large annual control program has been dedicating to reducing lamprey population abundance. Control measures are mainly based on construction of barriers at the mouths of Great Lakes rivers and through the use of lampricides such as 3-trifluormethlyl-4-nitrophenol (TFM). Current population assessment techniques in Great Lakes tributaries...
Show moreSea lamprey (Petromyzon marinus) are a harmful invasive species in the Great Lakes. Their invasion resulted in the decimation of native fish populations. A large annual control program has been dedicating to reducing lamprey population abundance. Control measures are mainly based on construction of barriers at the mouths of Great Lakes rivers and through the use of lampricides such as 3-trifluormethlyl-4-nitrophenol (TFM). Current population assessment techniques in Great Lakes tributaries include mark-recapture to quantify census size of sea lamprey spawning populations. Larval assessments characterize stream population abundance and size/age composition to prioritize streams for TFM treatment. We expanded traditional assessment methodologies by generating reconstructed pedigrees and estimates of effective breeding size (Nb) and minimum spawning size (Ns) of sea lamprey populations using single nucleotide polymorphism (SNP) genotypes developed from RAD-capture of larval sea lamprey collected during annual river assessments. In Chapter 1, we evaluated efficacy of barriers to adult upstream passage in three streams by assigning larvae to reconstructed pedigrees to estimate the number and effective number of spawning adults. In Chapter 2, we elucidated the effects of several sampling, biotic, and environmental factors on Nb and Ns using multi-locus SNP genotypes from larval populations from 18 streams across the Great Lakes. Correlations between mark-recapture estimates of adult abundance and Nb and Ns were also tested, and no significant correlation was found. Additional analyses were conducted to examine the effects of sample size, number of SNP loci, and true Nb on estimated Nb and Ns using simulated sea lamprey populations. As true Nb increased, different methods of estimating Nb and Ns showed different types and levels of bias, highlighting the need for multiple methods of estimating these parameters. Additionally, bias decreased across all estimates when sample size and SNP size were large. Overall, the analyses conducted provided unique insight into sea lamprey spawning populations and have potential as an annual assessment technique for evaluating both current control and future alternative controls.
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Title
THE GENOMIC BASIS FOR FITNESS AND ECOMORPHOLOGICAL VARIATION IN RECOVERING POPULATIONS OF LAKE TROUT (SALVELINUS NAMAYCUSH) IN THE GREAT LAKES
Creator
Smith, Seth Robert
Date
2021
Collection
Electronic Theses & Dissertations
Description
Here I describe the development of novel genomic resources that will be fundamental for advancing a new generation of genomic research on Lake Trout (Salvelinus namaycush) including a high-density linkage map, an annotated chromosome-anchored genome assembly, and three high-throughput genotyping panels. We used these resources to identify genomic regions exhibiting signals of adaptive divergence between Lake Trout hatchery strains, some of which were found to underlie differences in fitness ...
Show moreHere I describe the development of novel genomic resources that will be fundamental for advancing a new generation of genomic research on Lake Trout (Salvelinus namaycush) including a high-density linkage map, an annotated chromosome-anchored genome assembly, and three high-throughput genotyping panels. We used these resources to identify genomic regions exhibiting signals of adaptive divergence between Lake Trout hatchery strains, some of which were found to underlie differences in fitness (survival and reproduction) between strains in the contemporary Lake Huron environment. Loci associated with differences in fitness between Seneca and Great Lakes origin strains were localized using local ancestry inference and local ancestry outlier tests. By evaluating locus specific allelic contributions of the ancestral Seneca Lake and Great Lakes-derived hatchery strains to naturally-produced wild Lake Huron populations across the genomes of F2 wild born individuals, we were able to determine that a subset of 7 genomic regions contributed to differences in fitness between Seneca Lake and Great Lakes origin individuals during the re-emergence of wild populations in Lake Huron. We also identified 2 genomic regions where Great Lakes origin alleles were favored by selection, 4 regions associated with hybrid vigor, and 2 regions potentially associated with hybrid inferiority in recovering wild Lake Trout populations. We also found that some Seneca origin alleles are only favored by selection on certain genetic backgrounds. For instance, F2 hybrids between the Seneca strain and Lake Michigan origin strains have an excess of Seneca origin haplotypes on chromosome Sna11, while this is not the case in hybrids between Lake Superior and Seneca origin individuals. Collectively, these results indicate that elevated performance of the Seneca strain can at least be partially attributed to a number of adaptive alleles at a small subset of genes. Additionally, the fact that Great Lakes origin alleles were favored by selection at two loci indicates that native strains contain some variation that provides a fitness advantage in Lake Huron. These admixture outlier regions contained a significant excess of genes related to swimming behavior and negative regulation of vascular wound healing, which strongly suggests that differences in fitness between strains are due to behavioral and physiological factors associated with the ability to avoid and survive predation by Sea Lamprey. Additionally, we carried out two studies seeking to identify genetic variation associated with habitat occupancy and phenotypic variation in Lake Trout. First, we carried out a quantitative trait locus (QTL) mapping study in which we identified loci associated with length and condition related traits, skin pigmentation, and body shape. We produced a linkage map for Lake Trout as a prerequisite for this work. The information on locus order obtained from the linkage map was also critical to the assembly of the Lake Trout genome and recombination rate estimates obtained from the linkage map were critical to the goal of assigning haplotype segments to their hatchery strain of origin for wild-born Lake Huron individuals. This study also allowed us to determine the location of the Lake Trout sex determination locus, determine centromere locations, and characterize structural differences (i.e., chromosomal inversions and translocations) between Lake Trout and other salmonid species. Second, we performed a genome-wide scan for loci associated with ecomorphological divergence in Lake Superior Lake Trout (specifically between lean, siscowet, and humper forms), and identified numerous regions with abnormally high levels of divergence between forms. These loci likely underlie variation in traits that differentiate forms, as well as traits that contributed to reproductive isolation historically. For example, the genomic region most strongly associated with length and condition (from our QTL mapping study) was also associated with ecomorphological divergence in Lake Superior and this region also contains a putative chromosomal inversion. Interestingly, we find that hybridization primarily occurred between humpers and siscowets and humpers and leans immediately preceding a genetic homogenization event that occurred in the late 1990s or early 2000s. Using a collection of samples over a multi-decade time series collected from the Apostle Islands, we show that levels of hybridization with humpers increased substantially starting in the 1980s.
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Title
DEVELOPMENT OF FINE-MAPPING RESOURCES FOR INTERROGATION OF A YIELD IMPACTING QTL ON THE 2D CHROMOSOME IN A BREAD WHEAT (TRITICUM AESTIVUM) AND AEGILOPS TAUSCHII NESTED ASSOCIATION MAPPING POPULATION
Creator
Turkus, Jonathan Dubau
Date
2021
Collection
Electronic Theses & Dissertations
Description
Bread wheat (Triticum aestivum) is one of the dominant grain crops across the world. As a mainstay of diets across the world, there is a constant pressure by the breeders and geneticists to identify yield promoting loci. In previous work, significant SNP associations were found on the 2D chromosome (approximately at 23.5Mb and 25.2Mb) for yield variation in a genome-wide association study (GWAS) of the D-genome Nested Association Mapping population (DNAM), an advanced-backcross nested...
Show moreBread wheat (Triticum aestivum) is one of the dominant grain crops across the world. As a mainstay of diets across the world, there is a constant pressure by the breeders and geneticists to identify yield promoting loci. In previous work, significant SNP associations were found on the 2D chromosome (approximately at 23.5Mb and 25.2Mb) for yield variation in a genome-wide association study (GWAS) of the D-genome Nested Association Mapping population (DNAM), an advanced-backcross nested association mapping population of five Ae. tauschii lines into an elite bread wheat background. To identify the signal source more precisely, fine-mapping tools were created in this work. Seven Kompetitive Allele Specific PCR (KASP) primer sets were designed for SNP markers between 22Mb and 30Mb, six of which displayed codominant behavior and one dominant. The KASP primer sets were identified the U6718 BC1F1 subpopulation within the DNAM as segregating for bread wheat and Ae. tauschii alleles between 22Mb and 30Mb on 2D chromosome. Linkage maps were created for the seven D-genome chromosomes for the U6718 subpopulation. QTL analysis defined the QTL as being between 21.3cM to 28.3cM on the 2D linkage map and 23.3Mb to 30.3Mb on the physical map. Using the defined 2DS QTL and the KASP primer sets, a fine-mapping population was created consisting of five heterozygous inbred families fixed 16 unique recombination events across the 2DS QTL. These developed tools will prove instrumental tools towards refining the understanding of causal loci of this yield QTL.
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Title
USING THE REGULATION OF GA2-OXIDASE AS A MODEL TO UNDERSTAND GIBBERELLIN’S ROLES IN APPLE AND ARABIDOPSIS THALIANA
Creator
Zhang, Songwen
Date
2021
Collection
Electronic Theses & Dissertations
Description
In the model plant Arabidopsis thaliana (Arabidopsis), where the gibberellin (GA) signaling pathway has been best studied, GAs promote flowering. However, in many woody perennial plants such as apple (Malus x domestica Borkh.), GAs generally have a repressive role in flowering. The genetic and molecular mechanisms of GA-mediated flowering repression in apple and other plants remain unknown. In this study, we carried out transcriptional profiling of apple shoot apices during the anticipated...
Show moreIn the model plant Arabidopsis thaliana (Arabidopsis), where the gibberellin (GA) signaling pathway has been best studied, GAs promote flowering. However, in many woody perennial plants such as apple (Malus x domestica Borkh.), GAs generally have a repressive role in flowering. The genetic and molecular mechanisms of GA-mediated flowering repression in apple and other plants remain unknown. In this study, we carried out transcriptional profiling of apple shoot apices during the anticipated period of floral induction and examined how gene regulation is influenced by exogenous GAs. GA treatment resulted in downregulation of a diversity of genes participating in GA biosynthesis, transport and reception, and strong upregulation of the GA catabolic GA2-ox (GA2-oxidase) genes, suggesting GA homeostasis is mediated at multiple levels in these tissues. We also observed rapid and strong upregulation of both of two copies of a gene previously observed to inhibit flowering in apple, MdTFL1 (TERMINAL FLOWER 1), offering a potential explanation for the flowering-inhibitory effects of GA in apple. These results provide a context for investigating factors that may transduce the GA signal in apple and contribute to a preliminary genetic framework for the repression of flowering by GAs in a woody perennial plant. GA2ox expression and the enzyme activity determines concentrations of specific GAs in the right structure at the right time and contributes to enabling GAs to execute their function precisely. In return, their expression is under tight regulation by GA. Taking advantage of this tight link between GA2ox and GA activity, we aimed to develop regulation of GA2ox as a model to evaluate GA activity and function in both apple and Arabidopsis with a focus on floral induction. Through analysis of high-quality apple genomes and our transcriptional datasets, we identified 16 canonical GA2ox-like genes in the apple genome, representing eight pairs of homoeologous genes. By analyzing their spatial and developmental expression, we found that at least half of the genes were expressed in all the structures studied. In addition, individual members of homoeologous pairs displayed both overlapping and distinct expression patterns, suggesting function redundance and divergency. Finally, we found that two pairs of homoeologous genes (MdGA2ox1A, -1B, -2A and -2B) may be responsible for regulating GA activity in the shoot apex during floral induction. These findings provide biological and evolutionary insights into GA biology in apple, particularly in floral induction. In Arabidopsis, where GA2ox has been relatively well studied, we developed a genetic tool kit comprising both CRISPR/Cas-induced, transgene-free single knockout mutants and transcriptional and translational GUS fusion lines for the seven canonical Arabidopsis GA2ox genes (AtGA2ox1-AtGA2ox8). Mutations in the loss-of-function mutants have presumably disrupted function of both two functional domains of the GA2ox proteins. By comparing GUS activity in the GUS fusion lines, we found that the transcribed region is required for appropriate expression for several genes. This tool kit can be used for a wide spectrum of GA biology.
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Title
Characterization of cyclic-GMP-AMP signaling and novel gene networks in the Vibrio Seventh Pandemic Islands of El Tor Vibrio cholerae
Creator
Severin, Geoffrey Blaine-Graessley
Date
2020
Collection
Electronic Theses & Dissertations
Description
Vibrio cholerae is the causative agent of the diarrheal disease cholera, for which there have been seven pandemics. The first six pandemics (1817-1923) have been attributed to strains of the classical biotype while the seventh pandemic (1961- current) is being perpetuated by circulating strains of the El Tor biotype. It is hypothesized that El Tor's acquisition of two unique genomic islands of unknown origins, Vibrio Seventh Pandemic Island 1 & 2 (VSP-1 & 2), potentiated its displacement of...
Show moreVibrio cholerae is the causative agent of the diarrheal disease cholera, for which there have been seven pandemics. The first six pandemics (1817-1923) have been attributed to strains of the classical biotype while the seventh pandemic (1961- current) is being perpetuated by circulating strains of the El Tor biotype. It is hypothesized that El Tor's acquisition of two unique genomic islands of unknown origins, Vibrio Seventh Pandemic Island 1 & 2 (VSP-1 & 2), potentiated its displacement of classical strains in both environmental and clinical reservoirs prior to the seventh pandemic. Despite their connection to the pandemic evolution of the El Tor biotype and the likelihood they encode a wealth of novel biological functions the majority of the 303036 ORFs in the VSP islands have remained uncharacterized. The works presented in the thesis represent examples of our collective efforts to understand the unique traits afforded to the El Tor biotype by the VSP islands. .In 2012, ten years after the initial discovery of the VSP islands, the cyclic di-nucleotide synthase DncV was identified in VSP-1 and shown to synthesize the novel second messenger cyclic-GMP-AMP (cGAMP). Despite this significant discovery, cGAMP remained an orphan second messenger in bacteria for six years. In Chapter 2, I present our identification of the first cGAMP signaling network in bacteria by connecting the synthesis of cGAMP to the allosteric activation of the VSP-1 encoded patatin-like phospholipase vc0178, now named CapV. This finding helped catalyze the revelation that homologous cyclic-dinucleotide signaling networks contribute to bacteriophage-immunity and are distributed widely across the bacterial phyla.dncV and capV represent just two of the 103031 ORFs encoded in VSP-1, therefore we hypothesized that cGAMP signaling may extend to include additional genes within the island. Bioinformatic analysis of VSP island genes performed in collaboration with the laboratory of Eva Top at the University of Idaho predicted that dncV was likely to share a biological pathway with the VSP-1 encoded putative deoxycytidylate deaminase vc0175, we have now named DcdV. In Chapter 3, I describe our ongoing efforts to understand the biological function of DcdV and the discovery of its novel negative regulator DifV.The work presented in this thesis has made fundamental contribution to understanding the pandemic evolution of the El Tor biotype and the utility homologous genes afford to their bacterial host while providing a roadmap for further exploration of the biological role of the VSP-1 and VSP-2 islands in pandemic V. cholerae.
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Title
Genomics of Beta vulgaris crop types : insights into tap root development and storage characteristics
Creator
Galewski, Paul John
Date
2020
Collection
Electronic Theses & Dissertations
Description
Cultivated Beta vulgaris L. (beet) is a species complex composed of several distinct crop types developed for specific end uses. The crop types include sugar beet, fodder beet, table beet and leaf beet/chard. The evolution of each crop type appears to have resulted from interactions between selection, drift, gene flow, recombination, and the sorting of ancestral variation. Beets are generally heterozygous and contain self-incompatibility mechanisms. Therefore, reproducing and maintaining the...
Show moreCultivated Beta vulgaris L. (beet) is a species complex composed of several distinct crop types developed for specific end uses. The crop types include sugar beet, fodder beet, table beet and leaf beet/chard. The evolution of each crop type appears to have resulted from interactions between selection, drift, gene flow, recombination, and the sorting of ancestral variation. Beets are generally heterozygous and contain self-incompatibility mechanisms. Therefore, reproducing and maintaining the genetic constitution of a single individual for genetic and phenotypic analysis is a challenge. Beet populations are the fundamental unit of improvement and contain the evolutionary and adaptive potential of the species. This research used several approaches which explore the utility of pooled population genomic sequencing to survey the organization and distribution of genetic diversity within cultivated B. vulgaris lineages, and give context and clarity to the genetics underlying important agronomic characters.Whole genome sequence data was produced for important varieties and germplasm releases which represent the B. vulgaris crop type lineages. Using population genetic and statistical methods, relationships were determined between populations. Lineage-specific variation, or variation unique to specific crop types, was uncovered and used to quantify the level of support for these groups as discrete units. Allele frequency was able to differentiate between crop types using Principle Components Analysis (PCA), suggesting positive selection for end use was a major driver of crop type divergence. PCA carried out on a chromosome-by-chromosome basis showed the relative contributions of specific chromosomes to crop type diversification. Gene diversity (e.g., expected heterozygosity) and FST proved powerful indicators of selection along the chromosome at nucleotide resolution. In total, 12.13% of loci within the genome were differentiated with respect to crop type. Interestingly, this corresponds to levels of divergence observed in studies of incipient speciation. Differentiated regions, indicated by FST outliers, contained 472 genes, or 1.6% of the 24,255 genes predicted in the reference genome assembly. Respectively, sugar beet, table beet, fodder beet, and chard genomes contained 16, 283, 2, and 171 genes characterized as differentiated between crop types. Cryptic relationships were observed between crop types due to a high degree of genetic variation shared between crop type lineages. Specific instances of common ancestry, sorting of ancestral variation, and admixture and introgression were identified, which explain the degree of substructure observed between specific crop types.The content and organization of diversity in beet genomes reflects a complex history related to B. vulgaris crop type diversification. With the exception of chard, much of the species' historical selection has focused on the improvement of root characters (e.g., root enlargement, biomass, dry matter content, and sucrose concentration). As a result, major differences in root morphology and physiology can be observed between these lineages. Measures of root development and physiology between crop types were compared, and interestingly, much of the phenotypic variation partitioned between crop types corresponds to candidate genes identified from analyses of genome-wide variation using FST and 2pq. Admixture and introgression appear to have shared specific variation involved in the reduction of lateral roots (e.g., Root primordium defective 1), root enlargement (e.g., Brevis radix-like 4, putative NAC domain-containing protein 94, cytokinin dehydrogenase 3), and biomass accumulation (e.g., 6-phosphofructo-2-kinase). High relationship coefficients and high correlations in allele frequency for this variation were observed, indicating the genetic variation influencing these characters may have been derived from a single origin. The development of beet into an economically viable sugar crop required both an enlarged root and an increase sucrose concentration. Genes were identified that may explain these physiological changes within the root (e.g., decrease in water concentration, increase in dry matter content and increase in sucrose concentrations). These genes correspond to shared variation, distributed among crop types, as well as lineage-specific variation, restricted to sugar beet lineages. Integrating selection, drift, and admixture into a putative demographic history of beet provides evidence for the role of specific genes in the development of beet crop types and the expression of novel phenotypic characters.
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Title
Genome-wide association study reveals genes associated with mite recruitment phenotypes in the domesticated grapevine (vitis vinifera
Creator
LaPlante, Erika R.
Date
2020
Collection
Electronic Theses & Dissertations
Description
Plants in the grape genus Vitis have varying densities of trichomes and mite-domatia on their leaves, which facilitate the recruitment, retention, and reproduction of beneficial mites. By increasing the abundance of mites on grape leaves, these phenotypes promote a defense mutualism contributing to the control of grape pests and pathogens. Identification of the genes controlling these phenotypes would inform our understanding of the genetics underlying mite-plant mutualistic interactions and...
Show morePlants in the grape genus Vitis have varying densities of trichomes and mite-domatia on their leaves, which facilitate the recruitment, retention, and reproduction of beneficial mites. By increasing the abundance of mites on grape leaves, these phenotypes promote a defense mutualism contributing to the control of grape pests and pathogens. Identification of the genes controlling these phenotypes would inform our understanding of the genetics underlying mite-plant mutualistic interactions and could lead to breeding domesticated Vitis vinifera L. varieties that are naturally defended against pathogens. Little is known about the genetics underlying mite recruitment phenotypes in Vitis. We conducted a GWAS to determine the genetic architecture of mite recruitment traits in V. vinifera using 399 cultivars from a common garden diversity panel. We investigated eight traits previously established in the literature associated with an increase in beneficial mite abundance. We found single nucleotide polymorphisms (SNPs) significantly associated with each mite recruitment trait investigated. Corresponding gene annotations of SNP genetic coordinates revealed notable gene associations, including a trichome development gene, and a physiological defense response gene, suggesting these genetic regions may have a large impact on mediating mite-plant interactions in this species. Our findings are among the first to investigate which genes underly ecologically important mutualisms between plants and beneficial mites and suggest promising candidate genes for breeding and genetic editing to increase naturally occurring predator-based defenses in grapes.
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Title
Elucidating the genetic mechanisms of flowering and the repression of floral initiation by fruit in apple (Malus x domestica Borkh.)
Creator
Gottschalk, Christopher Charles
Date
2020
Collection
Electronic Theses & Dissertations
Description
Many tree fruit crops exhibit yearly cyclical fluctuations in flowering and fruiting, including apple (Malus × domestica Borkh.). This phenomenon, known as biennial bearing, is an intriguing biological problem, as well as a production limitation of many high-value apple cultivars. The current hypotheses to explain biennial bearing focus on the repressive effects of fruit on the initiation of floral primordia, which would develop into flowers the following year. This floral repressive response...
Show moreMany tree fruit crops exhibit yearly cyclical fluctuations in flowering and fruiting, including apple (Malus × domestica Borkh.). This phenomenon, known as biennial bearing, is an intriguing biological problem, as well as a production limitation of many high-value apple cultivars. The current hypotheses to explain biennial bearing focus on the repressive effects of fruit on the initiation of floral primordia, which would develop into flowers the following year. This floral repressive response could be the result of nutrient competition between the spur apex and the strong sink of the developing fruit, or flowering-inhibitory gibberellins (GAs) that are produced in the seeds of the developing fruit and exported to the spur apex. However, the molecular mechanism by which fruit load and/or GAs represses floral initiation is unknown.The first aim of my dissertation was to identify the genes involved in the floral initiation pathway. Utilizing transcripts assembled from a transcriptome of the biennial cultivar 'Honeycrisp' and the known flowering genes from Arabidopsis, I identified a comprehensive list of flowering-related homologs based on sequence homology, phylogenetic relationship, and syntenic organization. The second aim was to characterize the expression of the flowering-related homologs during the period of floral initiation under crop load conditions that either promote or repress flowering. Homologs of AGL24/SVP, AP1, FT, LFY, and SPLs were strongly expressed in apices of floral-induced trees, supporting their presumed role as floral promoters. In contrast, a homolog of the floral repressor TERMINAL FLOWER 1 (MdTFL1-2), was strongly up-regulated in apices of fruit-bearing, floral-repressed trees. Additionally, genes involved in the deactivation of bioactive GAs were strongly up-regulated in the floral-repressed trees. The timing of floral initiation is also known to vary between cultivars in apple. However, the diversity in flowering gene regulation that underlies this variation is unknown. The third aim of my dissertation was to investigate diversity of gene expression in six different apple species and cultivars. Generally, the expression profiles of the flowering-related genes were similar, suggesting a widely conserved mechanism. However, a few key genes involved in the regulation of floral initiation and development exhibited differential expression. For example, during the floral initiation period, five of the six genotypes were found to have differentially expressed MdTFL1s. This result implicates a potential role for the MdTFL1s in determining the timing of floral initiation in addition to a role in biennial bearing. Plant growth regulators (PGRs) offer the potential to control flowering and biennial bearing in established apple plantings. This final aim was to evaluate and improve the efficacy of foliar-applied GA regiments to repress flowering in apple. This study included the evaluation of application timings, GA formulations and concentrations, and cultivar-specificity. The most successful PGR trial used GA3 to inhibit flowering in 'Honeycrisp' when a strong return bloom was expected. As a result of the decreased flowering and fruit load, the harvested fruit in the year following GA application exhibited higher fruit-quality. The results from this collection of studies provide insight into the molecular control of flowering and biennial bearing while demonstrating a practical approach to managing flowering in a high-value apple cultivar.
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Title
Genetic variability and mapping of cooking time and sensory attributes in andean dry beans
Creator
Bassett, Amber Nichole
Date
2020
Collection
Electronic Theses & Dissertations
Description
Cooking time, flavor, and texture of dry beans (Phaseolus vulgaris L.) are valued by consumers but are not major considerations of dry bean breeding programs. The aim of this research is 1) to investigate mechanisms underlying fast cooking times of select genotypes, 2) to characterize the genetic control of cooking time, flavor, and texture of cooked beans in a diversity panel and a recombinant inbred line population, and 3) to evaluate how fast-cooking bean genotypes process into canned...
Show moreCooking time, flavor, and texture of dry beans (Phaseolus vulgaris L.) are valued by consumers but are not major considerations of dry bean breeding programs. The aim of this research is 1) to investigate mechanisms underlying fast cooking times of select genotypes, 2) to characterize the genetic control of cooking time, flavor, and texture of cooked beans in a diversity panel and a recombinant inbred line population, and 3) to evaluate how fast-cooking bean genotypes process into canned products. The genetic mechanism of fast cooking time was assessed via the physical and compositional seed characteristics in a set of 8 genotypes. Faster cooking beans had thinner cotyledon cell walls and seed coat layers and lower seed coat percentage, seed weight, and total and insoluble fiber. To identify genomic loci underlying cooking time, flavor, and texture, genome-wide association (GWA) and quantitative trait loci (QTL) mapping approaches were used with 430 lines of the Andean Diversity Panel and 242 yellow recombinant inbred lines. Sensory attributes included total flavor, beany, vegetative, earthy, starchy, sweet, and bitter intensity as well as seed coat perception and cotyledon texture. SNPs and QTL were identified for most of the attributes, with QTL for earthy intensity having the most phenotypic variation explained. In both populations, sweet and starchy intensity were positively correlated and associated via PCA, but other trait associations were minimal. A subset of lines from the RIL population were evaluated for canning quality following different retort processing durations. For fast-cooking lines, canning quality improved with reduced retort processing time, revealing a potential cost-saving benefit to the canning industry. This information lays a foundation for targeting fast cooking times and specific sensory profiles in breeding programs.
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Title
Identification and characterization of molecular targets for disease management in tree fruit pathogens
Creator
Peng, Jingyu
Date
2020
Collection
Electronic Theses & Dissertations
Description
Fire blight, caused by Erwinia amylovora, is a devastating bacterial disease threatening the worldwide production of pome fruit trees, including apple and pear. Within host xylem vessels, E. amylovora cells restrict water flow and cause wilting symptoms through formation of biofilms, that are matrix-enmeshed surface-attached microcolonies of bacterial cells. Biofilm matrix of E. amylovora is primarily composed of several exopolysaccharides (EPSs), including amylovora, levan, and, cellulose....
Show moreFire blight, caused by Erwinia amylovora, is a devastating bacterial disease threatening the worldwide production of pome fruit trees, including apple and pear. Within host xylem vessels, E. amylovora cells restrict water flow and cause wilting symptoms through formation of biofilms, that are matrix-enmeshed surface-attached microcolonies of bacterial cells. Biofilm matrix of E. amylovora is primarily composed of several exopolysaccharides (EPSs), including amylovora, levan, and, cellulose. The final step of biofilm development is dispersal, which allows dissemination of a subpopulation of biofilm cells to resume the planktonic mode of growth and consequentially cause systemic infection. In this work, we demonstrate that identified the Hfq-dependent small RNA (sRNA) RprA positively regulates amylovoran production, T3SS, and flagellar-dependent motility, and negatively affects levansucrase activity and cellulose production. We also identified the in vitro and in vivo conditions that activate RprA, and demonstrated that RprA activation leads to decreased formation of biofilms and promotes the dispersal movement of biofilm cells. This work supports the involvement of RprA in the systemic infection of E. amylovora during its pathogenesis.Bacterial toxin-antitoxin (TA) systems are small genetic loci composed of a proteinaceous toxin and a counteracting antitoxin. In this work, we identified and characterized a chromosomally encoded hok/sok-like type I TA system in Erwinia amylovora Ea1189. Ectopic overexpression of hok caused massive cell death in E. amylovora and its toxicity can be partially reversed through co-overexpression of the cognate sRNA sok. Phenotypic and transcriptomic examination of E. amylovora cells expressing hok at subtoxic levels demonstrated that hok causes membrane rupture and proton motive force dissipation, upregulates expression levels of ATP biosynthesis genes and consequently cellular ATP levels. Hok also positively affects phage shock protein genes to protect cells from further membrane damage. Taken together, the hok/sok TA system, besides being potentially self-toxic, appears to facilitate E. amylovora to manage various stress responses when the toxin gene is expressed in low levels.The succinate dehydrogenase inhibitor (SDHI) is a broad-spectrum fungicide class that has been widely utilized in agricultural fields. Blumeriella jaapii, the causative agent of cherry leaf spot (CLS), is the most important limiting factor for tart cherry production in the Midwestern United States. In the last decade, reduced efficacy in using SDHIs, i.e. boscalid, fluopyram, and fluxapyroxad, for management of CLS has been observed in many research and commercial orchard sites. Through whole-genome sequencing of B. jaapii using both PacBio long-reads and Illumina short-reads, we identified mutations in SdhB or SdhC that are correlative to resistance of this fungus to boscalid, fluopyram, and/or fluxapyroxad. SdhB and SdhC are components of the succinate dehydrogenase complex that contributes fungal respiration. In view of the widely conserved sequences of the SdhB and SdhC genes in phytopathogenic Ascomycete fungi, we expressed the wild-type or mutated alleles of the B. jaapii Sdh gene in the soybean white mold pathogen Sclerotinia sclerotiorum. We successfully validated the functions of the mutations in Sdh genes of B. jaapii in conferring resistance to several SDHIs examined in this study. The S. sclerotiorum heterologous expression system was also validated to be highly effective in characterizing the functions of other Sdh mutations of B. cinerea, C. homoeocarpa, and M. fructicola. The approach developed in this study can potentially be widely applied to interrogate SDHI fungicide resistance mechanisms in other phytopathogenic ascomycetes.
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