Wheat is the world’s third largest food crop, and is relied upon as a food source by millions of people. Securing the supply of wheat is a problem because it is susceptible to many biotic and abiotic factors that limit production. One such factor, sprouting of the grain in the head, because of untimely rainfall prior to harvest, is a substantial problem worldwide. Pre-harvest sprouting has a significant impact on wheat growers, who suffer considerable economic hardship as a result of yield loss during harvesting and subsequent downgrading of their sprouted crops. Wheat processors are also affected by this problem, because sprouted grain has significantly altered chemical properties, making it unsuitable for its intended purpose, and often rendering it suitable for animal consumption only. This study investigated mechanisms of dormancy, in the diploid wheat Triticum tauschii (Coss.) Schmalh., to assess their suitability for use in hexaploid (bread) wheat to prevent pre-harvest sprouting. A soluble germination inhibitor was found in the bracts (palea, lemma and glumes) surrounding the grain of T. tauschii. Fractionation of an aqueous extract from the bracts, by HPLC, identified vanillic acid as being likely to be involved in this inhibition. Further analysis of the extract also identified a strong anti-oxidant capacity, indicating that part of the inhibition of germination may arise from the prevention of oxygen reaching the embryo.

Pre-harvest sprouting (PHS) in bread wheat (<i>Triticum aestivum</i> L.) is one of the major abiotic constraints influencing the production of high quality grain. The flour milled from sprouted wheat grains has increased Ñ-amylase activity as compared to non-sprouted grain. PHS negatively affects the properties of flour with deleterious effects on bread and noodle quality. White-grained wheat is generally more susceptible to PHS damage than red-grained wheat. The objectives of this study were to identify a suitable method for phenotyping PHS resistance and to identify PHS resistance genomic regions and markers that could be used for marker-assisted selection in wheat improvement programs. A doubled haploid (DH) mapping population from a cross between two white-grained spring wheat genotypes, Argent (non-dormant) and W98616 (dormant) was used in this study. Forty DH lines (20 dormant and 20 non-dormant) were evaluated for germination frequency, Falling Number, and Ñ-amylase activity in dry and water-imbibed seeds and spikes. The germination test was the most reliable method for measurement of PHS resistance, whereas the Falling Number and Ñ-amylase activity in dry harvested seeds could not be correlated to dormancy levels. However, a positive association (r = 0.60***) was detected between germination frequency and Ñ-amylase activity in imbibed seeds. To identify the genomic regions associated with PHS resistance, a genetic linkage map with a total genome coverage of 2,577 cM was developed. The map was constructed from 913 scored markers (356 SSR, 290 AFLP, 258 DArT and 9 EST) with an average marker density of 3.7 cM/marker. Five genomic regions on chromosomes 1A, 3A, 4A, 7A and 7D were associated with PHS resistance by interval mapping and all regions were contributed by the dormant parent W98616. A total of 60 Canadian wheat cultivars and experimental lines were screened with three SSR markers, DuPw004, barc170 and wmc650, located under the major quantitative trait locus (QTL) on chromosome 4A. The SSR markers explained 60-75% of the total variation in germination frequency among different wheat genotypes. By using the DuPw004 marker in marker-assisted back crossing, the population size in the BC1F1 and BC2F1 generations were reduced by 41% and 59%, respectively. Thus, the 4A QTL markers have been proven useful for marker-assisted selection of PHS resistance for wheat improvement.

Pre-harvest sprouting (PHS) in bread wheat (<i>Triticum aestivum</i> L.) is one of the major abiotic constraints influencing the production of high quality grain. The flour milled from sprouted wheat grains has increased Ñ-amylase activity as compared to non-sprouted grain. PHS negatively affects the properties of flour with deleterious effects on bread and noodle quality. White-grained wheat is generally more susceptible to PHS damage than red-grained wheat. The objectives of this study were to identify a suitable method for phenotyping PHS resistance and to identify PHS resistance genomic regions and markers that could be used for marker-assisted selection in wheat improvement programs. A doubled haploid (DH) mapping population from a cross between two white-grained spring wheat genotypes, Argent (non-dormant) and W98616 (dormant) was used in this study. Forty DH lines (20 dormant and 20 non-dormant) were evaluated for germination frequency, Falling Number, and Ñ-amylase activity in dry and water-imbibed seeds and spikes. The germination test was the most reliable method for measurement of PHS resistance, whereas the Falling Number and Ñ-amylase activity in dry harvested seeds could not be correlated to dormancy levels. However, a positive association (r = 0.60***) was detected between germination frequency and Ñ-amylase activity in imbibed seeds. To identify the genomic regions associated with PHS resistance, a genetic linkage map with a total genome coverage of 2,577 cM was developed. The map was constructed from 913 scored markers (356 SSR, 290 AFLP, 258 DArT and 9 EST) with an average marker density of 3.7 cM/marker. Five genomic regions on chromosomes 1A, 3A, 4A, 7A and 7D were associated with PHS resistance by interval mapping and all regions were contributed by the dormant parent W98616. A total of 60 Canadian wheat cultivars and experimental lines were screened with three SSR markers, DuPw004, barc170 and wmc650, located under the major quantitative trait locus (QTL) on chromosome 4A. The SSR markers explained 60-75% of the total variation in germination frequency among different wheat genotypes. By using the DuPw004 marker in marker-assisted back crossing, the population size in the BC1F1 and BC2F1 generations were reduced by 41% and 59%, respectively. Thus, the 4A QTL markers have been proven useful for marker-assisted selection of PHS resistance for wheat improvement.

In malting barley, rapid germination is desirable and linked to end use quality. Modern malting varieties have been bred for low seed dormancy leading to issues with pre-harvest sprouting in wetter growing regions. To maintain malting capacity while minimizing germination on the maternal plant requires in-depth understanding of the genetic regulation of dormancy in malting barley. Currently, the major effect QTLs SD1 and SD2 have been shown to influence dormancy across multiple populations of barley, though the physiological mechanisms involved remain unclear. To search for novel genetic regions that influence primary dormancy, three mapping populations were assessed including two Canadian biparental populations (Synch and Legci) as well as a diversity panel sourced from multiple locations worldwide (ICARDA AM-14). The SD2 locus had a major effect in the Synch population while the SD1 locus had a major effect in the Legci population and neither SD1 nor SD2 were linked to dormancy in the diversity panel. Instead, 14 additional marker trait associations were identified in AM-14 suggesting that investigating a broader range of genetic regulation of dormancy outside of North American varieties may provide solutions to regulate this trait. Additional testing on SD1 revealed that variation at this locus did not affect ABA sensitivity during germination or GA or ABA-regulated gene expression during grain fill. Indeed, lines containing the non-dormant SD1 allele germinate at a similar rate as the dormant SD1 seeds when the glumella was removed from the embryo. This indicated that the effect of the alanine aminotransferase gene underlying the SD1 allele is dependent on physical restriction on the embryo or the hypoxic effects produced by the glumella. Imposing a hypoxic (5% oxygen) environment on exposed embryos revealed an association between non-dormancy at SD1 and reduced sensitivity to the suppressive effects of hypoxia on germination. This suggests that alanine aminotransferase regulates dormancy release during barley germination at least in part through regulation of the seed’s response to hypoxia.

Doctor of Philosophy / Department of Agronomy / Guihua Bai / Allan K. Fritz / Wheat pre-harvest sprouting (PHS), germination of physiologically matured grains in a wheat spike before harvesting, can cause significant reduction in grain yield and end-use quality. Many quantitative trait loci (QTL) for PHS resistance have been reported in different sources. To determine the genetic architecture of PHS resistance and its relationship with grain color (GC) in US hard winter wheat, a genome-wide association study (GWAS) on both PHS resistance and GC was conducted using in a panel of 185 U.S. elite breeding lines and cultivars and 90K wheat SNP arrrays. PHS resistance was assessed by evaluating sprouting rates in wheat spikes harvested from both greenhouse and field experiments. Thirteen QTLs for PHS resistance were identified on 11 chromosomes in at least two experiments, and the effects of these QTLs varied among different environments. The common QTLs for PHS resistance and GC were identified on the long arms of the chromosome 3A and 3D, indicating pleiotropic effect of the two QTLs. Significant QTLs were also detected on chromosome arms 3AS and 4AL, which were not related to GC, suggesting that it is possible to improve PHS resistance in white wheat. To identify markers closely linked to the 4AL QTL, genotyping-by-sequencing (GBS) technology was used to analyze a population of recombinant inbred lines (RILs) developed from a cross between two parents, “Tutoumai A” and “Siyang 936”, contrasting in 4AL QTL. Several closely linked GBS SNP markers to the 4AL QTL were identified and some of them were coverted to KASP for marker-assisted breeding. To investigate effects of the two non-GC related QTLs on 3AS and 4AL, both QTLs were transferered from “Tutoumai A” and “AUS1408” into a susceptible US hard winter wheat breeding line, NW97S186, through marker-assisted backcrossing using the gene marker TaPHS1 for 3AS QTL and a tightly linked KASP marker we developed for 4AL QTL. The 3AS QTL (TaPHS1) significantly interacted with environments and genetic backgrounds, whereas 4AL QTL (TaMKK3-A) interacted with environments only. The two QTLs showed additive effects on PHS resistance, indicating pyramiding these two QTLs can increase PHS resistance. To improve breeding selection efficiency, genomic prediction using genome-wide markers and marker-based prediction (MBP) using selected trait-linked markers were conducted in the association panel. Among the four genomic prediction methods evaluated, the ridge regression best linear unbiased prediction (rrBLUP) provides the best prediction among the tested methods (rrBLUP, BayesB, BayesC and BayesC0). However, MBP using 11 significant SNPs identified in the association study provides a better prediction than genomic prediction. Therefore, for traits that are controlled by a few major QTLs, MBP may be more effective than genomic selection.

Doctor of Philosophy / Department of Agronomy / Guihua Bai / Guorong Zhang / Wheat yield and quality is influenced by many abiotic and biotic environmental factors. Pre-harvest sprouting (PHS) occurs when physiologically matured spikes are exposed to wet field conditions before harvest, which results in seed germination and causes significant losses in yield and end-use quality. Wheat stripe rust is one of the most important biotic factors reducing grain yield and quality. To investigate the genetic basis of the resistance to PHS and stripe rust in hard white winter wheat cultivars Danby and Tiger and develop molecular markers for marker- assisted breeding, a double haploid (DH) population, derived from those two cultivars, was genotyped with simple sequence repeats (SSR) markers and simple nucleotide polymorphism (SNP) markers. This DH population was assessed for resistance to PHS and stripe rust in both greenhouse and field experiments. For PHS, one major resistant quantitative trait locus (QTL) was consistently detected on the short arm of chromosome 3A in all three experiments conducted and explained 21.6% to 41.0% of the phenotypic variation (PVE). This QTL is corresponding to a previously cloned gene, TaPHS1. A SNP in the promoter of TaPHS1 co- segregated with PHS resistance in this mapping population. Meanwhile, two other QTLs, Qphs.hwwg-3B.1 and Qphs.hwwg-5A.1, were consistently detected on the chromosome arms 3BS and 5AL in two experiments. These two QTLs showed significant additive effects with TaPHS1 in improving PHS resistance. For stripe rust, three major QTLs were consistently detected in four out of six environments for infection type (IT) or disease severity (DS). Two of them, QYr.hwwg-2AS1 and QYr.hwwg-4BL1, contributed by the Danby allele explained up to 28.4% of PVE for IT and 60.5% of PVE for DS. The third QTL, QYr.hwwg-3BS1, contributed by the Tiger allele, had PVE values up to 14.7% for IT and 22.9% for DS. QYr.hwwg-2AS1 and QYr.hwwg- 4BL1 are likely the same resistance genes reported previously on chromosome arms 2AS and 4BL. However, QYr.hwwg-3BS1 might be different from the reported gene cluster near the distal end of 3BS where Yr57, Yr4, Yr30 and Sr2 were located. Significant additive effects on reducing IT and DS were observed among these three major QTLs. In order to pyramid multiple QTLs in breeding, user-friendly Kompetitive allele specific PCR (KASP) markers were successfully developed for several QTLs identified in this study. The QTLs and their interactions found in this study together with those novel flanking KASP markers developed will be useful not only for understanding genetic mechanisms of PHS and stripe rust resistance but also for marker- assisted breeding to improve wheat resistance to PHS and stripe rust by gene pyramiding.

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