Genome-wide association study of stem rust resistance in a world collection of cultivated barley

Key message

QTL conferring a 14–40% reduction in adult plant stem rust severity to multiple races of Pgt were found on chromosome 5H and will be useful in barley breeding.

Abstract

Stem rust, caused by Puccinia graminis f. sp. tritici (Pgt) is an important disease of barley. The resistance gene Rpg1 has protected the crop against stem rust losses for over 70 years in North America, but is not effective against the African Pgt race TTKSK (and its variants) nor the domestic race QCCJB. To identify resistance to these Rpg1-virulent races, the Barley iCore Collection, held by the United States Department of Agriculture-Agricultural Research Service National Small Grains Collection was evaluated for adult plant resistance (APR) and seedling resistance to race TTKSK and APR to race QCCJB and the Pgt TTKSK composite of races TTKSK, TTKST, TTKTK, and TTKTT. Using a genome-wide association study approach based on 6224 single nucleotide polymorphic markers, seven significant loci for stem rust resistance were identified on chromosomes 1H, 2H, 3H, and 5H. The most significant markers detected were 11_11355 and SCRI_RS_177017 at 71–75 cM on chromosome 5H, conferring APR to QCCJB and TTKSK composite. Significant markers were also detected for TTKSK seedling resistance on chromosome 5H. All markers detected on 5H were independent of the rpg4/Rpg5 complex at 152–168 cM. This study verified the importance of the 11_11355 locus in conferring APR to races QCCJB and TTKSK and suggests that it may be effective against other races in the Ug99 lineage.

     

Regulatory hotspots are associated with plant gene expression under varying soil phosphorus supply in Brassica rapa

Gene expression is a quantitative trait that can be mapped genetically in structured populations to identify expression quantitative trait loci (eQTL). Genes and regulatory networks underlying complex traits can subsequently be inferred. Using a recently released genome sequence, we have defined cis- and trans-eQTL and their environmental response to low phosphorus (P) availability within a complex plant genome and found hotspots of trans-eQTL within the genome. Interval mapping, using P supply as a covariate, revealed 18,876 eQTL. trans-eQTL hotspots occurred on chromosomes A06 and A01 within Brassica rapa; these were enriched with P metabolism-related Gene Ontology terms (A06) as well as chloroplast- and photosynthesis-related terms (A01). We have also attributed heritability components to measures of gene expression across environments, allowing the identification of novel gene expression markers and gene expression changes associated with low P availability. Informative gene expression markers were used to map eQTL and P use efficiency-related QTL. Genes responsive to P supply had large environmental and heritable variance components. Regulatory loci and genes associated with P use efficiency identified through eQTL analysis are potential targets for further characterization and may have potential for crop improvement.     

Identification of markers linked to the race Ug99 effective stem rust resistance gene Sr28 in wheat (Triticum aestivum L.)

Wheat stem rust caused by Puccinia graminis f. sp. tritici can cause devastating yield losses in wheat. Over the past several decades, stem rust has been controlled worldwide through the use of genetic resistance. Stem rust race TTKSK (Ug99), first detected in Uganda in 1998, threatens global wheat production because of its unique virulence combination. As the majority of the currently grown cultivars and advanced breeding lines are susceptible to race TTKSK, sources of resistance need to be identified and characterized to facilitate their use in agriculture. South Dakota breeding line SD 1691 displayed resistance to race TTKSK in the international wheat stem rust nursery in Njoro, Kenya. Seedling screening of progeny derived from SD 1691 crossed to susceptible LMPG-6 indicated that a single resistance gene was present. Allelism and race-specificity tests indicated the stem rust resistance gene in SD 1691 was Sr28. The chromosome arm location of Sr28 was previously demonstrated to be 2BL. We identified molecular markers linked to Sr28 and validated this linkage in two additional populations. Common spring wheat cultivars in the central United States displayed allelic diversity for markers flanking Sr28. These markers could be used to select for Sr28 in breeding populations and for combining Sr28 with other stem rust resistance genes.     

Mapping quantitative and qualitative disease resistance genes in a doubled haploid population of barley (Hordeum vulgare)

Stripe rust, leaf rust, and Barley Yellow Dwarf Virus (BYDV) are important diseases of barley (Hordeum vulgare L). Using 94 doubled-haploid lines (DH) from the cross of Shyri x Galena, multiple disease phenotype datasets, and a 99-marker linkage map, we determined the number, genome location, and effects of genes conferring resistance to these diseases. We also mapped Resistance Gene Analog Polymorphism (RGAP) loci, based on degenerate motifs of cloned disease resistance genes, in the same population. Leaf rust resistance was determined by a single gene on chromosome 1 (7H). QTLs on chromosomes 2 (2H), 3 (3H), 5 (1H), and 6 (6H) were the principal determinants of resistance to stripe rust. Two- locus QTL interactions were significant determinants of resistance to this disease. Resistance to the MAV and PAV serotypes of BYDV was determined by coincident QTLs on chromosomes 1 (7H), 4 (4H), and 5 (1H). QTL interactions were not significant for BYDV resistance. The associations of molecular markers with qualitative and quantitative disease resistance loci will be a useful information for marker-assisted selection. Received: 2 February 1999 / Accepted: 30 December 1999     

Inheritance of resistance to Ug99 stem rust in wheat cultivar Norin 40 and genetic mapping of Sr42

Stem rust, caused by Puccinia graminis f. sp. tritici, is a devastating disease of wheat. The emergence of race TTKSK (Ug99) and new variants in Africa threatens wheat production worldwide. The best method of controlling stem rust is to deploy effective resistance genes in wheat cultivars. Few stem rust resistance (Sr) genes derived from the primary gene pool of wheat confer resistance to TTKSK. Norin 40, which carries Sr42, is resistant to TTKSK and variants TTKST and TTTSK. The goal of this study was to elucidate the inheritance of resistance to Ug99 in Norin 40 and map the Sr gene(s). A doubled haploid (DH) population of LMPG-6/Norin 40 was evaluated for resistance to the race TTKST. Segregation of 248 DH lines fitted a 1:1 ratio (χ (2) 1:1= 0.58, p = 0.45), indicating a single gene in Norin 40 conditioned resistance to Ug99. This was confirmed by an independent F(2:3) population also derived from the cross LMPG-6/Norin 40 where a 1:2:1 ratio (χ (2)1:2:1 = 0.69, p = 0.71) was observed following the inoculation with race TTKSK. Mapping with DNA markers located this gene to chromosome 6DS, the known location of Sr42. PCR marker FSD_RSA co-segregated with Sr42, and simple sequence repeat (SSR) marker BARC183 was closely linked (0.5 cM) to Sr42. A previous study found close linkage between FSD_RSA and SrCad, a temporarily designated gene that also confers resistance to Ug99, thus Sr42 may be the same gene or allelic. Marker FSD_RSA is suitable for marker-assisted selection (MAS) in wheat breeding programs to improve stem rust resistance, including Ug99.     

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.     

Association mapping and gene-gene interaction for stem rust resistance in CIMMYT spring wheat germplasm

The recent emergence of wheat stem rust Ug99 and evolution of new races within the lineage threatens global wheat production because they overcome widely deployed stem rust resistance (Sr) genes that had been effective for many years. To identify loci conferring adult plant resistance to races of Ug99 in wheat, we employed an association mapping approach for 276 current spring wheat breeding lines from the International Maize and Wheat Improvement Center (CIMMYT). Breeding lines were genotyped with Diversity Array Technology (DArT) and microsatellite markers. Phenotypic data was collected on these lines for stem rust race Ug99 resistance at the adult plant stage in the stem rust resistance screening nursery in Njoro, Kenya in seasons 2008, 2009 and 2010. Fifteen marker loci were found to be significantly associated with stem rust resistance. Several markers appeared to be linked to known Sr genes, while other significant markers were located in chromosome regions where no Sr genes have been previously reported. Most of these new loci colocalized with QTLs identified recently in different biparental populations. Using the same data and Q?+?K covariate matrices, we investigated the interactions among marker loci using linear regression models to calculate P values for pairwise marker interactions. Resistance marker loci including the Sr2 locus on 3BS and the wPt1859 locus on 7DL had significant interaction effects with other loci in the same chromosome arm and with markers on chromosome 6B. Other resistance marker loci had significant pairwise interactions with markers on different chromosomes. Based on these results, we propose that a complex network of gene-gene interactions is, in part, responsible for resistance to Ug99. Further investigation may provide insight for understanding mechanisms that contribute to this resistance gene network.     

Association mapping of stem rust race TTKSK resistance in US barley breeding germplasm

Key message

Loci conferring resistance to the highly virulent African stem rust race TTKSK were identified in advanced barley breeding germplasm and positioned to chromosomes 5H and 7H using an association mapping approach.

Abstract

African races of the stem rust pathogen (Puccinia graminis f. sp. tritici) are a serious threat to barley production worldwide because of their wide virulence. To discover and characterize resistance to African stem rust race TTKSK in US barley breeding germplasm, over 3,000 lines/cultivars were assessed for resistance at the seedling stage in the greenhouse and also the adult plant stage in the field in Kenya. Only 12 (0.3 %) and 64 (2.1 %) lines exhibited a resistance level comparable to the resistant control at the seedling and adult plant stage, respectively. To map quantitative trait loci (QTL) for resistance to race TTKSK, an association mapping approach was conducted, utilizing 3,072 single nucleotide polymorphism (SNP) markers. At the seedling stage, two neighboring SNP markers (0.8 cM apart) on chromosome 7H (11_21491 and 12_30528) were found significantly associated with resistance. The most significant one found was 12_30528; thus, the resistance QTL was named Rpg-qtl-7H-12_30528. At the adult plant stage, two SNP markers on chromosome 5H (11_11355 and 12_31427) were found significantly associated with resistance. This resistance QTL was named Rpg-qtl-5H-11_11355 for the most significant marker identified. Adult plant resistance is of paramount importance for stem rust. The marker associated with Rpg-qtl-5H-11_11355 for adult plant resistance explained only a small portion of the phenotypic variation (0.02); however, this QTL reduced disease severity up to 55.0 % under low disease pressure and up to 21.1 % under heavy disease pressure. SNP marker 11_11355 will be valuable for marker-assisted selection of adult plant stem rust resistance in barley breeding.     

Development of wheat lines carrying stem rust resistance gene <Emphasis Type="Italic">Sr39</Emphasis> with reduced <Emphasis Type="Italic">Aegilops speltoides</Emphasis> chromatin and simple PCR markers for marker-assisted selection

The use of major resistance genes is a cost-effective strategy for preventing stem rust epidemics in wheat crops. The stem rust resistance gene Sr39 provides resistance to all currently known pathotypes of Puccinia graminis f. sp. tritici (Pgt) including Ug99 (TTKSK) and was introgressed together with leaf rust resistance gene Lr35 conferring adult plant resistance to P. triticina (Pt), into wheat from Aegilops speltoides. It has not been used extensively in wheat breeding because of the presumed but as yet undocumented negative agronomic effects associated with Ae. speltoides chromatin. This investigation reports the production of a set of recombinants with shortened Ae. speltoides segments through induction of homoeologous recombination between the wheat and the Ae. speltoides chromosome. Simple PCR-based DNA markers were developed for resistant and susceptible genotypes (Sr39#22r and Sr39#50s) and validated across a set of recombinant lines and wheat cultivars. These markers will facilitate the pyramiding of ameliorated sources of Sr39 with other stem rust resistance genes that are effective against the Pgt pathotype TTKSK and its variants.     

RMo1 confers blast resistance in barley and is located within the complex of resistance genes containing Mla, a powdery mildew resistance gene

Isolates of Magnaporthe oryzae (the causal agent of rice blast disease) can infect a range of grass species, including barley. We report that barley Hordeum vulgare cv. Baronesse and an experimental line, BCD47, show a range of resistance reactions to infection with two rice blast isolates. The complete resistance of Baronesse to the isolate Ken 54-20 is controlled by a single dominant gene, designated RMo1. RMo1 mapped to the same linkage map position on chromosome 1H as the powdery mildew resistance locus Mla and an expressed sequence tag (k04320) that corresponds to the barley gene 711N16.16. A resistance quantitative trait locus (QTL), at which Baronesse contributed the resistance allele, to the isolate Ken 53-33 also mapped at the same position as RMo1. Synteny analysis revealed that a corresponding region on rice chromosome 5 includes the bacterial blight resistance gene xa5. These results indicate that a defined region on the short arm of barley chromosome 1H, including RMo1 and Mla, harbors genes conferring qualitative and quantitative resistance to multiple pathogens. The partial resistance of BCD47 to Ken53-33 is determined by alleles at three QTL, two of which coincide with the linkage map positions of the mildew resistance genes mlo and Mlf.     

Gene network inference via structural equation modeling in genetical genomics experiments

Our goal is gene network inference in genetical genomics or systems genetics experiments. For species where sequence information is available, we first perform expression quantitative trait locus (eQTL) mapping by jointly utilizing cis-, cis-trans-, and trans-regulation. After using local structural models to identify regulator-target pairs for each eQTL, we construct an encompassing directed network (EDN) by assembling all retained regulator-target relationships. The EDN has nodes corresponding to expressed genes and eQTL and directed edges from eQTL to cis-regulated target genes, from cis-regulated genes to cis-trans-regulated target genes, from trans-regulator genes to target genes, and from trans-eQTL to target genes. For network inference within the strongly constrained search space defined by the EDN, we propose structural equation modeling (SEM), because it can model cyclic networks and the EDN indeed contains feedback relationships. On the basis of a factorization of the likelihood and the constrained search space, our SEM algorithm infers networks involving several hundred genes and eQTL. Structure inference is based on a penalized likelihood ratio and an adaptation of Occam's window model selection. The SEM algorithm was evaluated using data simulated with nonlinear ordinary differential equations and known cyclic network topologies and was applied to a real yeast data set.     

Genetics and mapping of stem rust resistance to Ug99 in the wheat cultivar Webster

New races of wheat stem rust, namely TTKSK (Ug99) and its variants, pose a threat to wheat production in the regions where they are found. The accession of the wheat cultivar Webster (RL6201) maintained at the Cereal Research Centre in Winnipeg, Canada, shows resistance to TTKSK and other races of stem rust. The purpose of this study was to study the inheritance of seedling resistance to stem rust in RL6201 and genetically map the resistance genes using microsatellite (SSR) markers. A population was produced by crossing the stem rust susceptible line RL6071 with Webster. The F₂ and F₃ were tested with TPMK, a stem rust race native to North America. The F₃ was also tested with TTKSK. Two independently assorting genes were identified in RL6201. Resistance to TPMK was conferred by Sr30, which was mapped with microsatellites on chromosome 5DL. The second gene, temporarily designated SrWeb, conferred resistance to TTKSK. SrWeb was mapped to chromosome 2BL using SSR markers. Comparison with previous genetic maps showed that SrWeb occupies a locus near Sr9. Further analysis will be required to determine if SrWeb is a new gene or an allele of a previously identified gene.     

Quantitative trait loci controlling barley powdery mildew and scald resistances in two different barley doubled haploid populations

Powdery mildew and scald can cause significant yield loss in barley. In order to identify new resistance genes for powdery mildew and scald in barley, two barley doubled haploid (DH) populations were screened for adult plant resistance in the field and glasshouse under natural infection. The mapping populations included 92 DH lines from the cross of TX9425 × Franklin and 177 DH lines from the cross of Yerong × Franklin. Two quantitative trait loci (QTL) for resistance to powdery mildew were identified in the TX9425 × Franklin population. These QTL were mapped to chromosomes 7H and 5H, respectively. The phenotypic variation explained by the two QTL detected in this population was 22 and 17%, respectively. Three significant QTL were identified from the Yerong × Franklin population for the resistance to powdery mildew; the major one, detected on the short arm of chromosome 1H, explained 66% of phenotypic variation. The major QTL for scald resistance, identified from two different populations which shared a common parent, Franklin, were mapped in the similar position on 3H. However, the Franklin allele provided resistance to one population but susceptibility to the other population. The Yerong allele on 3H showed much better resistance to scald than the Franklin allele, which has not been reported before. Using high-density maps for both populations, some markers which were very close to the resistance genes were identified. Transgression beyond the parents in disease resistances of the DH populations indicates that both small-effect QTLs and genetic background may also have significant contributions towards the resistance.     

Cloning and characterisation of a family of disease resistance gene analogs from wheat and barley

 The most common class of plant disease resistance (R) genes cloned so far belong to the NBS-LRR group which contain nucleotide-binding sites (NBS) and a leucine-rich repeat (LRR). Specific primer sequences derived from a previously isolated NBS-LRR sequence at the Cre3 locus, which confers resistance to cereal cyst nematode (CCN) in wheat (Triticum aestivum L.) were used in isolating a family of resistance gene analogs (RGA) through a polymerase chain reaction (PCR) cloning approach. The cloning, analysis and genetic mapping of a family of RGAs from wheat (cv ‘Chinese Spring’) and barley (Hordeum vulgare L. cvs ‘Chebec’ and ‘Harrington’) are presented. The wheat and barley RGAs contain other conserved motifs present in known R genes from other plants and share between 55–99% amino acid sequence identity to the NBS-LRR sequence at the Cre3 locus. Phylogenetic analysis of the RGAs with other cloned R genes and RGAs from various plant species indicate that they belong to a superfamily of NBS-containing genes. Two of the barley derived RGAs were mapped onto loci on chromosomes 2H (2), 5H (7) and 7H (1) using barley doubled haploid (DH) mapping populations. Some of these loci identified are associated with regions carrying resistance to CCN and corn leaf aphid. Received: 6 January 1998 / Accepted: 1 April 1998