Glucosinolates are amino acid derived secondary metabolites found in the order Capparales. It is an important class of phytochemicals involved in plant-microbe, plant-insect, plant-animal and plant-human interactions. It is, therefore, important to understand genetic mechanism of glucosinolate biosynthesis in Brassica for efficient manipulation. In this study, QTL mapping of leaf and seed glucosinolates was performed in B. rapa using two RIL populations, SR-RILs and BU-RILs. QTL mapping was performed using SR-RILs developed from a cross of Chinese cabbage and turnip rapeseed and a genetic map in B.rapa. Genetic map was developed using a total 1,579 molecular markers including 9 markers specific to glucosinolate genes, GSL-ELONG, GSL-PRO, GSL-FMOOX1, and GSL-AOP/ALK. Several QTL for progoitrin, gluconapin, glucoalyssin, glucobrassicanapin, 2-methylpropyl and 4-hydoxyglucobrassicin glucosinolates were identified with phenotype variance between 6 and 54%. Interestingly, a major QTL for 5C aliphatic glucosinolates was co-localized with a candidate Br-GSL-ELONG locus on linkage group A3, displayed co-segregation with co-dominant SCAR marker BrMAM1-1. The Br-GSL-ELONG locus was identified to regulate 20 µmole/g seed 5C glucosinolate biosynthesis. BU-RILs derived from a cross of yellow sarson and USU9 was segregated for glucoerucin, gluconapin and progoitrin 4C aliphatic glucosinolates with 4-hydoxyglucobrassicin. Phenotyping was performed in controlled and field environments for seed glucosinolates and controlled environments for leaf glucosinolates. Genetic map was developed using SRAP markers and glucosinolate gene, GSL-ELONG and GSL-PRO specific 4 loci were integrated on map. Four and three QTL were identified for seed glucoerucin and gluconapin, respectively in both environments with phenotypic variance up to 49%. Additionally, genetic manipulation of glucosinolates was performed by backcross with MAS in B. rapa. Resynthesized B. napus line was backcrossed with B. rapa genotypes, RI16, BAR6 and USU9 for replacement or introgression of glucosinolate genes, GSL-ELONG- and GSL-PRO+. In RI16 genotype, 15 to 25 µmole/g seed 5C glucosinolates reduced in 15 BC3F2 lines those were positive with GSL-ELONG- marker and negative with the A-genome and gene specific marker BrMAM1-1. This suggests that the functional allele has replaced by non-functional from B. oleracea. GSL-PRO+ positive backcross lines in RI16 genotype displayed sinigrin 3C aliphatic glucosinolate in B. rapa. This suggests introgression of GSL-PRO+ in B. rapa.

Master of Science / Department of Plant Pathology / William Bockus / Robert Bowden / Stem rust, caused by the fungus Puccinia graminis f. sp. tritici, has historically been one of the most important diseases of wheat. Although losses have been much reduced in the last fifty years, new highly virulent races of the pathogen have recently emerged in East Africa. These new races are virulent on nearly all of the currently deployed resistance genes and therefore pose a serious threat to global wheat production. The spring wheat variety ‘Kingbird’ is thought to contain multiple quantitative trait loci (QTLs) that provide durable, adult-plant resistance against wheat stem rust. Stem rust-susceptible Kansas winter wheat line ‘KS05HW14’ was backcrossed to Kingbird and 379 recombinant lines were advanced to BC₁F₅ and then increased for testing. The lines were screened for stem rust resistance in the greenhouse and field in Kansas and in the field in Kenya over multiple years. We identified 16,237 single nucleotide polymorphisms (SNPs) with the Wheat 90K iSelect SNP Chip assay. After filtering for marker quality, linkage maps were constructed for each wheat chromosome. Composite interval mapping and multiple-QTL mapping identified seven QTLs on chromosome arms 2BL, 2DS, 3BS, 3BSc, 5DL, 7BL, and 7DS. Six QTLs were inherited from Kingbird and one QTL on 7BL was inherited from KS05HW14. The location of the QTL on 2BL is approximately at locus Sr9, 3BS is at Sr2, 3BSc is at Sr12, and 7DS is at Lr34/Yr18/Sr57. Although no QTL was found on 1BL, the presence of resistance gene Lr46/Yr29/Sr58 on 1BL in both parents was indicated by the gene-specific marker csLV46. QTLs on 2DS and 5DL may be related to photoperiod or vernalization genes. Pairwise interactions were only observed with race QFCSC, most notably occurring with QTLs 2BL and 3BSc. These results confirm that there are multiple QTLs present in Kingbird. Ultimately, the identification of the QTLs that make Kingbird resistant will aid in the understanding of durable, non-race-specific resistance to stem rust of wheat.

京都大学 / 新制・課程博士 / 博士(農学) / 甲第23618号 / 農博第2481号 / 新制||農||1088(附属図書館) / 学位論文||R4||N5366(農学部図書室) / 京都大学大学院農学研究科地域環境科学専攻 / (主査)教授 日本 典秀, 教授 田中 千尋, 准教授 刑部 正博 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

The goal of this thesis is to explore, improve and implement some advanced modern computational methods in statistics, focusing on applications in genetics. The thesis has three major directions. First, we study likelihoods for genetics analysis of experimental populations. Here, the maximum likelihood can be viewed as a computational global optimization problem. We introduce a faster optimization algorithm called PruneDIRECT, and explain how it can be parallelized for permutation testing using the Map-Reduce framework. We have implemented PruneDIRECT as an open source R package, and also Software as a Service for cloud infrastructures (QTLaaS). The second part of the thesis focusses on using sparse matrix methods for solving linear mixed models with large correlation matrices. For populations with known pedigrees, we show that the inverse of covariance matrix is sparse. We describe how to use this sparsity to develop a new method to maximize the likelihood and calculate the variance components. In the final part of the thesis we study computational challenges of psychiatric genetics, using only pedigree information. The aim is to investigate existence of maternal effects in obsessive compulsive behavior. We add the maternal effects to the linear mixed model, used in the second part of this thesis, and we describe the computational challenges of working with binary traits. / eSSENCE

Doctor of Philosophy / Department of Agronomy / Jianming Yu / Knowledge of the genetic bases of grain quality traits will complement plant breeding efforts to improve the end use value of sorghum (Sorghum bicolor (L.) Moench). The objective of the first experiment was to assess marker-trait associations for 10 grain quality traits through candidate gene association mapping on a diverse panel of 300 sorghum accessions. The 10 grain quality traits were measured using the single kernel characterization system (SKCS) and near-infrared reflectance spectroscopy (NIRS). The analysis of the accessions through 1,290 genome-wide single nucleotide polymorphisms (SNPs) separated the panel into five subpopulations that corresponded to three major sorghum races (durra, kafir, and caudatum), one intermediate race (guinea-caudatum), and one working group (zerazera/caudatum). Association analysis between 333 SNPs in candidate genes/loci and grain quality traits resulted in eight significant marker-trait associations. A SNP in starch synthase IIa (SSIIa) gene was associated with kernel hardness (KH) with a likelihood ratio–based R[superscript]2 (R[subscript]L[subscript]R[superscript]2) value of 0.08. SNPs in starch synthase (SSIIb) gene (R[subscript]L[subscript]R[superscript]2 = 0.10) and loci pSB1120 (R[subscript]L[subscript]R[superscript]2 = 0.09) was associated with starch content. Sorghum is a crop well adapted to the semi arid regions of the world and my harbor genes for drought tolerance. The objective of second experiment was to identify quantitative trait loci (QTLs) for yield potential and drought tolerance. From a cross between Tx436 (food grain type) and 00MN7645 (drought tolerant) 248 recombinant inbred lines (RILs) was developed. Multi-location trials were conducted in 8 environments to evaluate agronomic performance of the RILs under favorable and drought stress conditions. The 248 RILs and their parents were genotyped by genotyping-by-sequencing (GBS). A subset of 800 SNPs was used for linkage map construction and QTL detection. Composite interval mapping identified a major QTLs for grain yield in chromosome 8 and QTL for flowering time in chromosome 9 under favorable conditions. Three major QTLs were detected for grain yield in chromosomes 1, 6, and 8 and two flowering time QTLs on chromosome 1 under drought conditions. Six QTLs were identified for stay green: two on chromosome 4; one each on chromosome 5, 6, 7, and 10 under drought conditions.

Master of Science / Department of Agronomy / Allan Fritz / Fusarium head blight (FHB) is one of the most devastative diseases in wheat. Growing resistant cultivars is one of the most effective strategies to minimize the disease damage. Huangcandou (HCD) is a Chinese wheat landrace showing a high level of resistance to FHB spread within a spike (type II). To identify quantitative traits loci (QTL) for resistance in HCD, a population of 190 recombinant inbred lines (RILs) were developed from a cross between HCD and Jagger, a susceptible hard winter wheat (HWW) released in Kansas. The population was evaluated for type II resistance at the greenhouses of Kansas State University. After initial marker screening, 261 polymorphic simple-sequence repeats (SSR) between parents were used for analysis of the RIL population. Among three QTL identified, two from HCD were mapped on the short arms of chromosomes 3B (3BS) and 3A (3AS). The QTL on the distal end of 3BS showed a major effect on type II resistance in all three experiments. This QTL coincides with a previously reported Fhb1, and explained 28.3% of phenotypic variation. The QTL on 3AS explained 9.7% of phenotypic variation for mean PSS over three experiments. The third QTL from chromosome 2D of Jagger explained 6.5% of phenotypic variation. Allelic substitution using the closest marker to each QTL revealed that substitution of Jagger alleles of two QTL on 3AS and 3BS with those from HCD significantly reduced the PSS. HCD containing both QTL on 3AS and 3BS with a large effect on type II resistance can be an alternative source of FHB resistance for improving FHB type II resistance in wheat. Besides, meta-analyses were used to estimate 95% confidence intervals (CIs) of 24 mapped QTL in five previously mapped populations derived from Chinese landraces: Wangshuibai (WSB), Haiyanzhong (HYZ), Huangfangzhu (HFZ), Baishanyuehuang (BSYH) and Huangcandou (HCD). Nineteen QTL for FHB type II resistance were projected to 10 QTL clusters. Five QTL on chromosomes 1A, 5A, 7A, and 3BS (2) were identified as confirmed QTL that have stable and consistent effects on FHB resistance and markers in these meta-QTL regions should be useful for marker-assisted breeding.

The incidence of infectious diseases in livestock is a major concern for animal breeders as well as for consumers. As a alternative approach to the use of prophylactic measures or therapeutic agents, infectious diseases can be contended by increasing the disease resistance of animals by genetic improvement. Animals can be selected either on a measure of their resistance (indicator trait) or on the presence or absence of some specific resistance genes in their genotype. A prerequisite to the latter approach is the identification of the genes, or QTL for quantitative trait loci, underlying the trait of interest. By means of sophisticated statistical tools, the QTL mapping strategy combines the information from genetic markers and phenotypic values to dissect quantitative traits into their individual genetic components. Some of the methodological aspects of this strategy are studied in the present thesis in the context of disease resistance in livestock. Indicator traits of the resistance (such as bacteria or parasites counts) are not always satisfying the normality assumption underlying most of the QTL mapping methods. In this context, the ability of statistical tests to identify the underlying genes (i.e. the statistical power) can be considerably reduced. We show that compared to the use of a non-parametric method, the use of the least-squares-based parametric method on mathematically transformed phenotypes gives always the best results. In the context of high number of ties (equal values) as observed when measuring resistance to bacterial or parasitic diseases, the non-parametric test is a good alternative to this approach, as far as midranks are used for ties instead of random ranks. The efficiency of QTL mapping methods can also be increased by use of simple combinations of repeated measurements of the same trait. As a result of analyses performed on real data sets in chicken and sheep, we show that much attention should be paid to obtaining good quality measurements, reflecting at best differences in terms of resistance between animals, before performing a QTL search. The appropriate choice of resistance traits as well as of the time of their measurement are, beside the choice of the method and the quality of marker information, among the most preponderant factors to guarantee satisfying results.

Lately the rate of genetic gain in most agronomic crop species has been reduced due to several factors that limit breeding efficiency and genetic gain. New genetic tools and more powerful statistical analyses provide an alternative approach to enhance genetic improvements through the identification of molecular markers linked to genomic regions or QTLs controlling quantitative traits. The main objective of this research was to identify genomic regions associated with enhanced agronomic performance in lines per se and hybrid combination in Sorghum bicolor (L.) Moench. A population composed of 187 F5:6 recombinant inbred lines (RIL) was derived from the cross of restorer lines RTx430 and RTx7000. Also, a testcross hybrid population (TCH) was developed by using each RIL as a pollinator onto ATx2752. A linkage map was constructed using 174 marker loci generated from AFLP and SSR primer combinations. These markers were assigned to 12 different linkage groups. The linkage map covers 1573 cM with marker loci spaced at an averaged 9.04 cM. In this study, 89 QTL that control variation in seven different morphological traits were identified in the recombinant inbred line population, while in the testcross hybrid population, 79 QTL were identified. These traits included grain yield, plant height, days to mid-anthesis, panicle number, panicle length, panicle exsertion and panicle weight. These putative QTL explained from 4 to 42% of the phenotypic variation observed for each trait. Many of the QTL were not consistent across populations and across environments. Nevertheless, a few key QTL were identified and the source of the positive additive genetics isolated. RTx7000 was consistently associated with better agronomic performance in RIL, while in testcrosses, RTx430 was. Some genomic regions from RTx7000 may be utilized to improve RTx430 as a line per se. However, it is very unlikely that such regions will have a positive effect on the combining ability of RTx430 since testcross results did not reveal any transgressive segregants from the RIL population.

Lately the rate of genetic gain in most agronomic crop species has been reduced due to several factors that limit breeding efficiency and genetic gain. New genetic tools and more powerful statistical analyses provide an alternative approach to enhance genetic improvements through the identification of molecular markers linked to genomic regions or QTLs controlling quantitative traits. The main objective of this research was to identify genomic regions associated with enhanced agronomic performance in lines per se and hybrid combination in Sorghum bicolor (L.) Moench. A population composed of 187 F5:6 recombinant inbred lines (RIL) was derived from the cross of restorer lines RTx430 and RTx7000. Also, a testcross hybrid population (TCH) was developed by using each RIL as a pollinator onto ATx2752. A linkage map was constructed using 174 marker loci generated from AFLP and SSR primer combinations. These markers were assigned to 12 different linkage groups. The linkage map covers 1573 cM with marker loci spaced at an averaged 9.04 cM. In this study, 89 QTL that control variation in seven different morphological traits were identified in the recombinant inbred line population, while in the testcross hybrid population, 79 QTL were identified. These traits included grain yield, plant height, days to mid-anthesis, panicle number, panicle length, panicle exsertion and panicle weight. These putative QTL explained from 4 to 42% of the phenotypic variation observed for each trait. Many of the QTL were not consistent across populations and across environments. Nevertheless, a few key QTL were identified and the source of the positive additive genetics isolated. RTx7000 was consistently associated with better agronomic performance in RIL, while in testcrosses, RTx430 was. Some genomic regions from RTx7000 may be utilized to improve RTx430 as a line per se. However, it is very unlikely that such regions will have a positive effect on the combining ability of RTx430 since testcross results did not reveal any transgressive segregants from the RIL population.

The cactophilic Drosophila mojavensis species group living in the deserts and dry tropical forests of the Southwestern United States and Mexico provides a valuable system for studies in diversification and speciation. My dissertation addresses a variety of evolutionary genetic questions using this system.Rigorous studies of the relationships between host races of D. mojavensis and the relationships among the members of the species group (D. mojavensis, D. arizona, and D. navojoa) are lacking. I used mitochondrial CO1 sequence data to address the phylogenetics and population genetics of this species group (Appendix A). In this study I have found that the sister species D. mojavensis and D. arizonae share no mitochondrial haplotypes and thus show no evidence for recent introgression. I estimate the divergence time between D. mojavensis and D. arizonae to be between 0.66 and 0.99 million years ago. I performed additional population genetic analyses of these species to provide a basis for future hypothesis testing.In Appendix B, I report the first example of substantial intraspecific polymorphism for genetic factors contributing to hybrid male sterility. I show that the occurrence of hybrid male sterility in crosses between Drosophila mojavensis and its sister species, D. arizonae is controlled by factors present at different frequencies in different populations of D. mojavensis. In addition, I show that hybrid male sterility is a complex phenotype; some hybrid males with motile sperm still cannot sire offspring.The large degree of variation between isofemale lines in producing sterile hybrid sons suggests a complex genetic basis to hybrid male sterility warranting quantitative genetic analysis. Since the genes underlying hybrid male sterility in these species are not yet fixed, I am able to perform explicit genetic analysis of this reproductive isolating mechanism. In Appendix C, I present the results of mapping QTL for hybrid male sterility within species. The genetic architecture underlying hybrid male sterility when analyzed directly in the F1 is highly complex. Thus, hybrid male sterility arises as a complex trait in this system and we propose a drift-based model for the evolution of this phenotype.