Validation of Molecular Markers for Use With Adapted Sources of Fusarium Head Blight Resistance in Wheat.

Genetic control of resistance to Fusarium head blight (FHB) is quantitative, making phenotypic selection difficult. Genetic markers to resistance are helpful to select favorable genotypes. This study was conducted to determine if Fhb1 and Fhb5 present in the Sumai 3 source of FHB resistance occur in Sumai 3-derived North American spring wheat cultivars and to understand the appropriateness of using markers to select for the favorable alleles at these loci in breeding. Sumai 3-derived parents Alsen, ND3085, ND744, Carberry, and Glenn were used in crosses to generate 14 doubled haploid breeding populations. The parents and progeny were genotyped with five Fhb1 and three Fhb5 microsatellite markers. Progeny were selected based on performance relative to parents and other control cultivars in FHB nurseries near Portage la Prairie and Carman, MB. χ2 and t test analyses were performed on marker and FHB data. The χ2 test frequently determined the proportion of lines carrying molecular variants associated with FHB resistance increased following nursery selection for FHB. Similarly, the t test regularly demonstrated that selection for FHB resistance lowered the mean level of disease associated with resistant marker haplotypes. The study affirmed FHB resistance sources Alsen, Carberry, ND3085, and ND744 have Fhb1 and Fhb5 loci like Sumai 3, but no evidence was found that Glenn carries Fhb1 and Fhb5 resistance alleles. The results justified use of Fhb1 and Fhb5 markers for marker assisted selection in populations derived from Alsen, Carberry, ND3085, and ND744, but not Glenn. Combined or individual application of Xgwm493 and Xgwm533 in selection of genotypes carrying Fhb1, and Xgwm150, Xgwm304, and Xgwm595 for Fhb5 will enhance FHB resistance in wheat.

Potential to breed for mycorrhizal association in durum wheat.

The selection of genotypes under high soil fertility may alter the effectiveness of mycorrhizal symbioses naturally forming between crop plants and the mycorrhizal fungi residing in cultivated fields. We tested the hypothesis that the mycorrhizal symbiosis of 5 landraces functions better than the mycorrhizal symbiosis of 27 cultivars of durum wheat that were bred after the development of the fertilizer industry. We examined the development of mycorrhiza and the response of these genotypes to mycorrhiza formation after 4 weeks of growth under high and low soil fertility levels in the greenhouse. The durum wheat genotypes were seeded in an established extraradical hyphal network of Rhizophagus irregularis and in a control soil free of mycorrhizal fungi. The percentage of root length colonized by mycorrhizal fungi was lower in landraces (21%) than in cultivars (27%; P = 0.04) and in the most recent releases (29%; P = 0.02), which were selected under high soil fertility levels. Plant growth response to mycorrhiza varied from -36% to +19%. Overall, durum wheat plant breeding in Canada has increased the mycorrhizal development in wheat grown at a low soil fertility level. However, breeding had inconsistent effects on mycorrhizal development and has led to the production of cultivars with patterns of regulation ranging from unimproved to inefficient.

Stripe rust and leaf rust resistance QTL mapping, epistatic interactions, and co-localization with stem rust resistance loci in spring wheat evaluated over three continents.

KEY MESSAGE: In wheat, advantageous gene-rich or pleiotropic regions for stripe, leaf, and stem rust and epistatic interactions between rust resistance loci should be accounted for in plant breeding strategies. Leaf rust (Puccinia triticina Eriks.) and stripe rust (Puccinia striiformis f. tritici Eriks) contribute to major production losses in many regions worldwide. The objectives of this research were to identify and study epistatic interactions of quantitative trait loci (QTL) for stripe and leaf rust resistance in a doubled haploid (DH) population derived from the cross of Canadian wheat cultivars, AC Cadillac and Carberry. The relationship of leaf and stripe rust resistance QTL that co-located with stem rust resistance QTL previously mapped in this population was also investigated. The Carberry/AC Cadillac population was genotyped with DArT(®) and simple sequence repeat markers. The parents and population were phenotyped for stripe rust severity and infection response in field rust nurseries in Kenya (Njoro), Canada (Swift Current), and New Zealand (Lincoln); and for leaf rust severity and infection response in field nurseries in Canada (Swift Current) and New Zealand (Lincoln). AC Cadillac was a source of stripe rust resistance QTL on chromosomes 2A, 2B, 3A, 3B, 5B, and 7B; and Carberry was a source of resistance on chromosomes 2B, 4B, and 7A. AC Cadillac contributed QTL for resistance to leaf rust on chromosome 2A and Carberry contributed QTL on chromosomes 2B and 4B. Stripe rust resistance QTL co-localized with previously reported stem rust resistance QTL on 2B, 3B, and 7B, while leaf rust resistance QTL co-localized with 4B stem rust resistance QTL. Several epistatic interactions were identified both for stripe and leaf rust resistance QTL. We have identified useful combinations of genetic loci with main and epistatic effects. Multiple disease resistance regions identified on chromosomes 2A, 2B, 3B, 4B, 5B, and 7B are prime candidates for further investigation and validation of their broad resistance.

Genetics of pre-harvest sprouting resistance in a cross of Canadian adapted durum wheat genotypes.

Severe losses attributable to pre-harvest sprouting (PHS) have been reported in Canada in recent years. The genetics of PHS resistance have been more extensively studied in hexaploid wheat and generally not using combinations of elite agronomic parents. The objective of our research was to understand the genetic nature of PHS resistance in an elite durum cross. A doubled haploid (DH) population and checks were phenotyped in replicated trials for grain yield and PHS traits over 3 years in western Canada. The response of intact spikes to sprouting conditions, sampled over two development time points, was measured in a rain simulation chamber. The DH population was genotyped with simple sequence repeat and Diversity Arrays Technology markers. Genotypes were a significant source of variation for grain yield and PHS resistance traits in each tested environment. Transgressive segregant DH genotypes were identified for grain yield and PHS resistance measurements. Low or no correlation was detected between grain yield and PHS, while correlation between PHS resistance measurements was moderate. The heritability of PHS resistance was moderate and higher than grain yield. Significant quantitative trait loci with small effect were detected on chromosomes 1A, 1B, 5B, 7A and 7B. Both parents contributed to the PHS resistance. Promising DH genotypes with high and stable grain yield as well as PHS resistance were identified, suggesting that grain yield and PHS can be improved simultaneously in elite genetic materials, and that these DH genotypes will be useful parental material for durum breeding programs.

Identification and mapping in spring wheat of genetic factors controlling stem rust resistance and the study of their epistatic interactions across multiple environments.

Stem rust (Puccinia graminis f. sp. tritici) is responsible for major production losses in hexaploid wheat (Triticum aestivum L.) around the world. The spread of stem rust race Ug99 and variants is a threat to worldwide wheat production and efforts are ongoing to identify and incorporate resistance. The objectives of this research were to identify quantitative trait loci (QTL) and to study their epistatic interactions for stem rust resistance in a population derived from the Canadian wheat cultivars AC Cadillac and Carberry. A doubled haploid (DH) population was developed and genotyped with DArT(®) and SSR markers. The parents and DH lines were phenotyped for stem rust severity and infection response to Ug99 and variant races in 2009, 2010 and 2011 in field rust nurseries near Njoro, Kenya, and to North American races in 2011 and 2012 near Swift Current, SK, Canada. Seedling infection type to race TTKSK was assessed in a bio-containment facility in 2009 and 2012 near Morden, MB. Eight QTL for stem rust resistance and three QTL for pseudo-black chaff on nine wheat chromosomes were identified. The phenotypic variance (PV) explained by the stem rust resistance QTL ranged from 2.4 to 48.8 %. AC Cadillac contributed stem rust resistance QTL on chromosomes 2B, 3B, 5B, 6D, 7B and 7D. Carberry contributed resistance QTL on 4B and 5A. Epistatic interactions were observed between loci on 4B and 5B, 4B and 7B, 6D and 3B, 6D and 5B, and 6D and 7B. The stem rust resistance locus on 6D interacted synergistically with 5B to improve the disease resistance through both crossover and non-crossover interactions depending on the environment. Results from this study will assist in planning breeding for stem rust resistance by maximizing QTL main effects and epistatic interactions.

Identification and mapping of leaf, stem and stripe rust resistance quantitative trait loci and their interactions in durum wheat.

Leaf rust (Puccinia triticina Eriks.), stripe rust (Puccinia striiformis f. tritici Eriks.) and stem rust (Puccinia graminis f. sp. tritici) cause major production losses in durum wheat (Triticum turgidum L. var. durum). The objective of this research was to identify and map leaf, stripe and stem rust resistance loci from the French cultivar Sachem and Canadian cultivar Strongfield. A doubled haploid population from Sachem/Strongfield and parents were phenotyped for seedling reaction to leaf rust races BBG/BN and BBG/BP and adult plant response was determined in three field rust nurseries near El Batan, Obregon and Toluca, Mexico. Stripe rust response was recorded in 2009 and 2011 nurseries near Toluca and near Njoro, Kenya in 2010. Response to stem rust was recorded in field nurseries near Njoro, Kenya, in 2010 and 2011. Sachem was resistant to leaf, stripe and stem rust. A major leaf rust quantitative trait locus (QTL) was identified on chromosome 7B at Xgwm146 in Sachem. In the same region on 7B, a stripe rust QTL was identified in Strongfield. Leaf and stripe rust QTL around DArT marker wPt3451 were identified on chromosome 1B. On chromosome 2B, a significant leaf rust QTL was detected conferred by Strongfield, and at the same QTL, a Yr gene derived from Sachem conferred resistance. Significant stem rust resistance QTL were detected on chromosome 4B. Consistent interactions among loci for resistance to each rust type across nurseries were detected, especially for leaf rust QTL on 7B. Sachem and Strongfield offer useful sources of rust resistance genes for durum rust breeding.

Genetic variability in arbuscular mycorrhizal fungi compatibility supports the selection of durum wheat genotypes for enhancing soil ecological services and cropping systems in Canada.

Crop nutrient- and water-use efficiency could be improved by using crop varieties highly compatible with arbuscular mycorrhizal fungi (AMF). Two greenhouse experiments demonstrated the presence of genetic variability for this trait in modern durum wheat ( Triticum turgidum L. var. durum Desf.) germplasm. Among the five cultivars tested, 'AC Morse' had consistently low levels of AM root colonization and DT710 had consistently high levels of AM root colonization, whereas 'Commander', which had the highest colonization levels under low soil fertility conditions, developed poor colonization levels under medium fertility level. The presence of genetic variability in durum wheat compatibility with AMF was further evidenced by significant genotype × inoculation interaction effects in grain and straw biomass production; grain P, straw P, and straw K concentrations under medium soil fertility level; and straw K and grain Fe concentrations at low soil fertility. Mycorrhizal dependency was an undesirable trait of 'Mongibello', which showed poor growth and nutrient balance in the absence of AMF. An AMF-mediated reduction in grain Cd under low soil fertility indicated that breeding durum wheat for compatibility with AMF could help reduce grain Cd concentration in durum wheat. Durum wheat genotypes should be selected for compatibility with AMF rather than for mycorrhizal dependency.

Targeted mapping of Cdu1, a major locus regulating grain cadmium concentration in durum wheat (Triticum turgidum L. var durum).

Some durum wheat (Triticum turgidum L. var durum) cultivars have the genetic propensity to accumulate cadmium (Cd) in the grain. A major gene controlling grain Cd concentration designated as Cdu1 has been reported on 5B, but the genetic factor(s) conferring the low Cd phenotype are currently unknown. The objectives of this study were to saturate the chromosomal region harboring Cdu1 with newly developed PCR-based markers and to investigate the colinearity of this wheat chromosomal region with rice (Oryza sativa L.) and Brachypodium distachyon genomes. Genetic mapping of markers linked to Cdu1 in a population of recombinant inbred substitution lines revealed that the gene(s) associated with variation in Cd concentration resides in wheat bin 5BL9 between fraction breakpoints 0.76 and 0.79. Genetic mapping and quantitative trait locus (QTL) analysis of grain Cd concentration was performed in 155 doubled haploid lines from the cross W9262-260D3 (low Cd) by Kofa (high Cd) revealed two expressed sequence tag markers (ESMs) and one sequence tagged site (STS) marker that co-segregated with Cdu1 and explained >80% of the phenotypic variation in grain Cd concentration. A second, minor QTL for grain Cd concentration was also identified on 5B, 67 cM proximal to Cdu1. The Cdu1 interval spans 286 kbp of rice chromosome 3 and 282 kbp of Brachypodium chromosome 1. The markers and rice and Brachypodium colinearity described here represent tools that will assist in the positional cloning of Cdu1 and can be used to select for low Cd accumulation in durum wheat breeding programs targeting this trait. The isolation of Cdu1 will further our knowledge of Cd accumulation in cereals as well as metal accumulation in general.

Chromosomal location of the cadmium uptake gene (Cdu1) in durum wheat.

Levels of the heavy metal cadmium (Cd) in food products are a food safety concern. Grain Cd is higher in durum (Triticum turgidum L. var. durum) than in common wheat, so reduction of Cd in durum grain is a priority of breeding programs. Previous research demonstrated that a single dominant gene, Cdu1, confers the low grain Cd phenotype, but the map location of the gene is not known. A doubled haploid population segregating for Cd concentration, developed from the cross of W9262-260D3 (a Kyle*2/Biodur inbred selection with low Cd uptake) and Kofa (high Cd uptake) and mapped with microsatellite markers, was used to locate Cdu1. Grain Cd concentration was determined by standard laboratory methods on field grain samples in 2000 and 2001. The Cd concentration segregated bimodally, allowing Cdu1 to be mapped qualitatively as well as quantitatively with quantitative trait locus analysis. The Cdu1 gene mapped to the long arm of chromosome 5B.

Identification and validation of quantitative trait loci for grain protein concentration in adapted Canadian durum wheat populations.

Grain protein concentration (GPC) is one of the most important factors influencing pasta-making quality. Durum wheat (Triticum turgidum L. var durum) cultivars with high GPC produce pasta with increased tolerance to overcooking and greater cooked firmness. However, the large environmental effect on expression of GPC and the negative correlation with grain yield have slowed genetic improvement of this important trait. Understanding the genetics and identification of molecular markers associated with high GPC would aid durum wheat breeders in trait selection at earlier generations. The objectives of this study were to identify and validate molecular markers associated with quantitative trait loci (QTL) for elevated GPC in durum wheat. A genetic map was constructed using SSR and DArT markers in an F(1)-derived doubled haploid (DH) population derived from the cross DT695 x Strongfield. The GPC data were collected from replicated trials grown in six Canadian environments from 2002 to 2005. QTL associated with variation for GPC were identified on the group 1, 2, and 7 chromosomes and on 5B and 6B, but only QGpc.usw-B3 on 2B and QGpc.usw-A3 on 7A were expressed consistently in four and six environments, respectively. Positive alleles for GPC at these loci were contributed by the high-GPC parent Strongfield. The QGpc.usw-A3 QTL was validated in a second DH population, and depending on environment, selection for the Strongfield allele at barc108 resulted in +0.4% to +1.0% increase in GPC, with little effect on yield in most environments. Given the consistent expression pattern in multiple populations and environments, barc108 could be useful for marker-assisted selection for high GPC.

Allelic variation at Psy1-A1 and association with yellow pigment in durum wheat grain.

The yellow pigment (YP) of durum wheat (Triticum turgidum L. var durum) semolina is due in part to the presence of carotenoid pigments found in the endosperm and is an important end-use quality trait. Phytoene synthase (Psy) is considered a rate-limiting enzyme in the carotenoid biosynthetic pathway and in this study, three alleles of Psy1-A1 were sequenced from four durum wheat cultivars and a co-dominant marker was developed for genetic mapping. Psy1-A1 mapped to chromosome 7AL near Xwmc809 in three durum mapping populations and was significantly associated with a pigment quantitative trait loci (QTL) identified on that chromosome. A second QTL localized 25 cM proximal to Psy1-A1 in two populations, and the interaction between the two QTL was not significant. Consistent with QTL mapping data, the Psy1-A1o allele was associated with elevated pigment in a validation population comprising 93 diverse cultivars and breeding lines. These results confirm an earlier hypothesis that Psy1, and at least one additional gene in the distal region of 7AL, are associated with grain YP differences in durum wheat. The functional co-dominant marker developed in this study differentiates the Psy1-A1 alleles reported here and could be used as a target to enhance YP selection in durum wheat breeding programs.

Association mapping of yellow pigment in an elite collection of durum wheat cultivars and breeding lines.

Association mapping (AM) is an alternative or complementary strategy to QTL mapping for describing associations between genotype and phenotype based on linkage disequilibrium (LD). Yellow pigment (YP), an important end-use quality trait in durum wheat (Triticum turgidum L. var. durum), was evaluated to determine the ability of AM to identify previously published QTL and to identify genomic regions for further genetic dissection. The YP concentration was determined for 93 durum wheat accessions sampled from a variety of geographic origins. Analysis of population structure using distance- and model-based estimates indicated the presence of five subpopulations. Using subpopulation assignments as covariates, significant (P < 0.05) marker-trait associations for YP were detected on all chromosomes of the durum genome. Using AM, genomic regions housing known YP QTL were confirmed, most notably the group 7 chromosomes. In addition, several markers on the group 1, 2, and 3 chromosomes were identified where QTL have yet to be reported. A phytoene synthase gene, Psy1-B1, a potential candidate gene for YP, was significantly associated with YP and was in strong LD with microsatellite markers on the distal end of 7BL. Our results demonstrated that AM complemented traditional QTL mapping techniques and identified novel QTL that should be the target of further genetic dissection.

Identification of QTL and association of a phytoene synthase gene with endosperm colour in durum wheat.

The yellow colour of durum wheat (Triticum turgidum L. var durum) semolina is due in part to the presence of carotenoid pigments found in the endosperm and is an important end-use quality trait. We hypothesized that variation in the genes coding for phytoene synthase (Psy), a critical enzyme in carotenoid biosynthesis, may partially explain the phenotypic variation in endosperm colour observed among durum cultivars. Using rice sequence information, primers were designed to PCR clone and sequence the Psy genes from Kofa (high colour) and W9262-260D3 (medium colour) durum cultivars. Sequencing confirmed the presence of four Psy genes in each parent, corresponding to a two member gene family designated as Psy1-1, Psy1-2 and Psy2-1 and Psy2-2. A genetic map was constructed using 155 F1-derived doubled haploid lines from the cross W9262-260D3/Kofa with 194 simple sequence repeat and DArT markers. Using Psy1-1 and Psy2-1 allele-specific markers and chromosome mapping, the Psy1 and Psy2 genes were located to the group 7 and 5 chromosomes, respectively. Four quantitative trait loci (QTL) underlying phenotypic variation in endosperm colour were identified on chromosomes 2A, 4B, 6B, and 7B. The Psy1-1 locus co-segregated with the 7B QTL, demonstrating an association of this gene with phenotypic variation for endosperm colour. This work is the first report of mapping Psy genes and supports the role of Psy1-1 in elevated levels of endosperm colour in durum wheat. This gene is a target for the further development of a molecular marker to enhance selection for endosperm colour in durum wheat breeding programs.

Black Point Reaction of Durum and Common Wheat Cultivars Grown Under Irrigation in Southern Saskatchewan.

Six durum (Triticum turgidum L. var. durum) and six common (T. aestivum L.) wheat cultivars were compared for reaction to black point under irrigation at two locations in southern Saskatchewan in 1990 to 1992 and 1994. There were individual varietal differences in black point levels within each of the species. The Canada Western Soft White Spring wheat Fielder was the most susceptible and the Canada Western Red Spring wheat Katepwa was the most resistant to black point. The location-cultivar-year interaction was a significant source of variation and a crossover cultivar-environment interaction was significant, suggesting that rank order of cultivars differed with environment. The durum wheat had significantly higher black point levels than the common wheat cultivars in three of the seven environments conducive to black point, two of which were in 1992, and had high overall black point levels. This greater black point severity on the durum wheat cultivars might have been due to cool, wet weather conditions and frosts during seed development that delayed ripening.

DNA markers linked to a T10 loose smut resistance gene in wheat (Triticum aestivum L.).

Screening for loose smut resistance in wheat is difficult. Selecting lines with DNA markers linked to loose smut resistance would be more reliable and less costly. Molecular markers linked to a race T10 loose smut resistance gene were identified using a F6 single seed descent segregating population. A RAPD marker and a RFLP marker were located on opposite flanks of the resistance gene and were shown to be loosely linked. The RAPD marker was converted to a user friendly polymorphic SCAR marker that represented a single genetically defined locus in hexaploid wheat. Using these two bracketing markers simultaneously, the error rate for T10 resistance selection due to crossing-over was reduced to 4%. These markers can be used for a faster and more reliable selection of T10 resistant plants than previous conventional loose smut ratings.

A monoclonal antibody chromosome marker analysis used to locate a loose smut resistance gene in wheat chromosome 6A.

Many genes have been located in wheat chromosomes, yet little is known about the location of genes for resistance to Ustilago tritici, which causes loose smut. Crosses were made between the loose smut susceptible alien substitution lines Cadet 6Ag(6A) and Rescue 6Ag(6A) (lines in which Agropyron chromosome 6 is substituted by wheat chromosome 6A) and four cultivars resistant to U. tritici race T19: 'Cadet', 'Kota', 'Thatcher' and 'TD18'. The segregating progeny were tested for reaction to race T19 and for the level of binding with a monoclonal antibody specific to a chromosome 6A-coded seed protein. The antibody, which does not bind to seed protein extracts in the absence of the 6A chromosome, was used as a chromosome marker. An association was established between resistance to race T19 and the presence of chromosome 6A for each of the cultivars tested, indicating that resistance to race T19 resides in chromosome 6A. Ustilago tritici race T19 resistance in 'Cadet' appears to be located in the short arm of chromosome 6A, based on the evaluation of the Cadet 6A long ditelosomic stock, which was susceptible, and the Cadet 6A-short: 6-Agropyron- short alien translocation stock, which was resistant.