Dissection of a rapidly evolving wheat resistance gene cluster by long-read genome sequencing accelerated the cloning of Pm69.

Gene cloning in repeat-rich polyploid genomes remains challenging. Here, we describe a strategy for overcoming major bottlenecks in cloning of the powdery mildew resistance gene (R-gene) Pm69 derived from tetraploid wild emmer wheat. A conventional positional cloning approach was not effective owing to suppressed recombination. Chromosome sorting was compromised by insufficient purity. A Pm69 physical map, constructed by assembling Oxford Nanopore Technology (ONT) long-read genome sequences, revealed a rapidly evolving nucleotide-binding leucine-rich repeat (NLR) R-gene cluster with structural variations. A single candidate NLR was identified by anchoring RNA sequencing reads from susceptible mutants to ONT contigs and was validated by virus-induced gene silencing. Pm69 is likely a newly evolved NLR and was discovered in only one location across the wild emmer wheat distribution range in Israel. Pm69 was successfully introgressed into cultivated wheat, and a diagnostic molecular marker was used to accelerate its deployment and pyramiding with other R-genes.

Genome scans capture key adaptation and historical hybridization signatures in tetraploid wheat.

Tetraploid wheats (Triticum turgidum L.), including durum wheat (T. turgidum ssp. durum (Desf.) Husn.), are important crops with high nutritional and cultural values. However, their production is constrained by sensitivity to environmental conditions. In search of adaptive genetic signatures tracing historical selection and hybridization events, we performed genome scans on two datasets: (1) Durum Global Diversity Panel comprising a total of 442 tetraploid wheat and wild progenitor accessions including durum landraces (n = 286), domesticated emmer (T. turgidum ssp. dicoccum (Schrank) Thell.; n = 103) and wild emmer (T. turgidum ssp. dicoccoides (Korn. ex Asch. & Graebn.) Thell.; n = 53) wheats genotyped using the 90K single nucleotide polymorphism (SNP) array, and (2) a second dataset comprising a total 121 accessions of nine T. turgidum subspecies including wild emmer genotyped with >100 M SNPs from whole-genome resequencing. The genome scan on the first dataset detected six outlier loci on chromosomes 1A, 1B, 3A (n = 2), 6A, and 7A. These loci harbored important genes for adaptation to abiotic stresses, phenological responses, such as seed dormancy, circadian clock, flowering time, and key yield-related traits, including pleiotropic genes, such as HAT1, KUODA1, CBL1, and ZFN1. The scan on the second dataset captured a highly differentiated region on chromosome 2B that shows significant differentiation between two groups: one group consists of Georgian (T. turgidum ssp. paleocolchicum A. Love & D. Love) and Persian (T. turgidum ssp. carthlicum (Nevski) A. Love & D. Love) wheat accessions, while the other group comprises all the remaining tetraploids including wild emmer. This is consistent with a previously reported introgression in this genomic region from T. timopheevii Zhuk. which naturally cohabit in the Georgian and neighboring areas. This region harbored several adaptive genes, including the thermomorphogenesis gene PIF4, which confers temperature-resilient disease resistance and regulates other biological processes. Genome scans can be used to fast-track germplasm housed in gene banks and in situ; which helps to identify environmentally resilient accessions for breeding and/or to prioritize them for conservation.

Multi-locus genome-wide association studies reveal the genetic architecture of Fusarium head blight resistance in durum wheat.

Durum wheat is more susceptible to Fusarium head blight (FHB) than other types or classes of wheat. The disease is one of the most devastating in wheat; it reduces yield and end-use quality and contaminates the grain with fungal mycotoxins such as deoxynivalenol (DON). A panel of 265 Canadian and European durum wheat cultivars, as well as breeding and experimental lines, were tested in artificially inoculated field environments (2019-2022, inclusive) and two greenhouse trials (2019 and 2020). The trials were assessed for FHB severity and incidence, visual rating index, Fusarium-damaged kernels, DON accumulation, anthesis or heading date, maturity date, and plant height. In addition, yellow pigment and protein content were analyzed for the 2020 field season. To capture loci underlying FHB resistance and related traits, GWAS was performed using single-locus and several multi-locus models, employing 13,504 SNPs. Thirty-one QTL significantly associated with one or more FHB-related traits were identified, of which nine were consistent across environments and associated with multiple FHB-related traits. Although many of the QTL were identified in regions previously reported to affect FHB, the QTL QFhb-3B.2, associated with FHB severity, incidence, and DON accumulation, appears to be novel. We developed KASP markers for six FHB-associated QTL that were consistently detected across multiple environments and validated them on the Global Durum Panel (GDP). Analysis of allelic diversity and the frequencies of these revealed that the lines in the GDP harbor between zero and six resistance alleles. This study provides a comprehensive assessment of the genetic basis of FHB resistance and DON accumulation in durum wheat. Accessions with multiple favorable alleles were identified and will be useful genetic resources to improve FHB resistance in durum breeding programs through marker-assisted recurrent selection and gene stacking.

Historical Selection, Adaptation Signatures, and Ambiguity of Introgressions in Wheat.

Wheat was one of the crops domesticated in the Fertile Crescent region approximately 10,000 years ago. Despite undergoing recent polyploidization, hull-to-free-thresh transition events, and domestication bottlenecks, wheat is now grown in over 130 countries and accounts for a quarter of the world's cereal production. The main reason for its widespread success is its broad genetic diversity that allows it to thrive in different environments. To trace historical selection and hybridization signatures, genome scans were performed on two datasets: approximately 113K SNPs from 921 predominantly bread wheat accessions and approximately 110K SNPs from about 400 wheat accessions representing all ploidy levels. To identify environmental factors associated with the loci, a genome-environment association (GEA) was also performed. The genome scans on both datasets identified a highly differentiated region on chromosome 4A where accessions in the first dataset were dichotomized into a group (n = 691), comprising nearly all cultivars, wild emmer, and most landraces, and a second group (n = 230), dominated by landraces and spelt accessions. The grouping of cultivars is likely linked to their potential ancestor, bread wheat cv. Norin-10. The 4A region harbored important genes involved in adaptations to environmental conditions. The GEA detected loci associated with latitude and temperature. The genetic signatures detected in this study provide insight into the historical selection and hybridization events in the wheat genome that shaped its current genetic structure and facilitated its success in a wide spectrum of environmental conditions. The genome scans and GEA approaches applied in this study can help in screening the germplasm housed in gene banks for breeding, and for conservation purposes.

The effect of wheat genotype on the microbiome is more evident in roots and varies through time.

Crop breeding has traditionally ignored the plant-associated microbial communities. Considering the interactions between plant genotype and associated microbiota is of value since different genotypes of the same crop often harbor distinct microbial communities which can influence the plant phenotype. However, recent studies have reported contrasting results, which led us to hypothesize that the effect of genotype is constrained by growth stages, sampling year and plant compartment. To test this hypothesis, we sampled bulk soil, rhizosphere soil and roots of 10 field-grown wheat genotypes, twice per year, for 4 years. DNA was extracted and regions of the bacterial 16 S rRNA and CPN60 genes and the fungal ITS region were amplified and sequenced. The effect of genotype was highly contingent on the time of sampling and on the plant compartment sampled. Only for a few sampling dates, were the microbial communities significantly different across genotypes. The effect of genotype was most often significant for root microbial communities. The three marker genes used provided a highly coherent picture of the effect of genotype. Taken together, our results confirm that microbial communities in the plant environment strongly vary across compartments, growth stages, and years, and that this can mask the effect of genotype.

Genetic mapping of leaf rust (Puccinia triticina Eriks) resistance genes in six Canadian spring wheat cultivars.

The Canada Western Red Spring wheat (Triticum aestivum L.) cultivars AAC Concord, AAC Prevail, CDC Hughes, Lillian, Glenlea, and elite line BW961 express a spectrum of resistance to leaf rust caused by Puccinia triticina Eriks. This study aimed to identify and map the leaf rust resistance of the cultivars using three doubled haploid populations, AAC Prevail/BW961 (PB), CDC Hughes/AAC Concord (HC), and Lillian/Glenlea (LG). The populations were evaluated for seedling resistance in the greenhouse and adult plant disease response in the field at Morden, MB for 3 years and genotyped with the 90K wheat Infinium iSelect SNP array. Genetic maps were constructed to perform QTL analysis on the seedling and field leaf rust data. A total of three field leaf rust resistance QTL segregated in the PB population, five in the HC, and six in the LG population. In the PB population, BW961 contributed two QTL on chromosomes 2DS and 7DS, and AAC Prevail contributed a QTL on 4AL consistent across trials. Of the five QTL in HC, AAC Concord contributed two QTL on 4AL and 7AL consistent across trials and a QTL on 3DL.1 that provided seedling resistance only. CDC Hughes contributed two QTL on 1DS and 3DL.2. Lillian contributed four QTL significant in at least two of the three trials on 2BS, 4AL, 5AL, and 7AL, and Glenlea two QTL on 4BL and 7BL. The 1DS QTL from CDC Hughes, the 2DS from BW961, the 4AL from the AAC Prevail, AAC Concord, and Lillian, and the 7AL from AAC Concord and Lillian conferred seedling leaf rust resistance. The QTL on 4AL corresponded with Lr30 and was the same across cultivars AAC Prevail, AAC Concord, and Lillian, whereas the 7AL corresponding with LrCen was coincident between AAC Concord and Lillian. The 7DS and 2DS QTL in BW961 corresponded with Lr34 and Lr2a, respectively, and the 1DS QTL in CDC Hughes with Lr21. The QTL identified on 5AL could represent a novel gene. The results of this study will widen our knowledge of leaf rust resistance genes in Canadian wheat and their utilization in resistance breeding.

Genetic architecture of rust resistance in a wheat (Triticum turgidum) diversity panel.

Introduction: Wheat rust diseases are widespread and affect all wheat growing areas around the globe. Breeding strategies focus on incorporating genetic disease resistance. However, pathogens can quickly evolve and overcome the resistance genes deployed in commercial cultivars, creating a constant need for identifying new sources of resistance. Methods: We have assembled a diverse tetraploid wheat panel comprised of 447 accessions of three Triticum turgidum subspecies and performed a genome-wide association study (GWAS) for resistance to wheat stem, stripe, and leaf rusts. The panel was genotyped with the 90K Wheat iSelect single nucleotide polymorphism (SNP) array and subsequent filtering resulted in a set of 6,410 non-redundant SNP markers with known physical positions. Results: Population structure and phylogenetic analyses revealed that the diversity panel could be divided into three subpopulations based on phylogenetic/geographic relatedness. Marker-trait associations (MTAs) were detected for two stem rust, two stripe rust and one leaf rust resistance loci. Of them, three MTAs coincide with the known rust resistance genes Sr13, Yr15 and Yr67, while the other two may harbor undescribed resistance genes. Discussion: The tetraploid wheat diversity panel, developed and characterized herein, captures wide geographic origins, genetic diversity, and evolutionary history since domestication making it a useful community resource for mapping of other agronomically important traits and for conducting evolutionary studies.

The coordinated regulation of early meiotic stages is dominated by non-coding RNAs and stage-specific transcription in wheat.

Reproductive success hinges on precisely coordinated meiosis, yet our understanding of how structural rearrangements of chromatin and phase transitions during meiosis are transcriptionally regulated is limited. In crop plants, detailed analysis of the meiotic transcriptome could identify regulatory genes and epigenetic regulators that can be targeted to increase recombination rates and broaden genetic variation, as well as provide a resource for comparison among eukaryotes of different taxa to answer outstanding questions about meiosis. We conducted a meiotic stage-specific analysis of messenger RNA (mRNA), small non-coding RNA (sncRNA), and long intervening/intergenic non-coding RNA (lincRNA) in wheat (Triticum aestivum L.) and revealed novel mechanisms of meiotic transcriptional regulation and meiosis-specific transcripts. Amidst general repression of mRNA expression, significant enrichment of ncRNAs was identified during prophase I relative to vegetative cells. The core meiotic transcriptome was comprised of 9309 meiosis-specific transcripts, 48 134 previously unannotated meiotic transcripts, and many known and novel ncRNAs differentially expressed at specific stages. The abundant meiotic sncRNAs controlled the reprogramming of central metabolic pathways by targeting genes involved in photosynthesis, glycolysis, hormone biosynthesis, and cellular homeostasis, and lincRNAs enhanced the expression of nearby genes. Alternative splicing was not evident in this polyploid species, but isoforms were switched at phase transitions. The novel, stage-specific regulatory controls uncovered here challenge the conventional understanding of this crucial biological process and provide a new resource of requisite knowledge for those aiming to directly modulate meiosis to improve crop plants. The wheat meiosis transcriptome dataset can be queried for genes of interest using an eFP browser located at https://bar.utoronto.ca/efp_wheat/cgi-bin/efpWeb.cgi?dataSource=Wheat_Meiosis.

A technical guide to TRITEX, a computational pipeline for chromosome-scale sequence assembly of plant genomes.

BACKGROUND: As complete and accurate genome sequences are becoming easier to obtain, more researchers wish to get one or more of them to support their research endeavors. Reliable and well-documented sequence assembly workflows find use in reference or pangenome projects. RESULTS: We describe modifications to the TRITEX genome assembly workflow motivated by the rise of fast and easy long-read contig assembly of inbred plant genomes and the routine deployment of the toolchains in pangenome projects. New features include the use as surrogates of or complements to dense genetic maps and the introduction of user-editable tables to make the curation of contig placements easier and more intuitive. CONCLUSION: Even maximally contiguous sequence assemblies of the telomere-to-telomere sort, and to a yet greater extent, the fragmented kind require validation, correction, and comparison to reference standards. As pangenomics is burgeoning, these tasks are bound to become more widespread and TRITEX is one tool to get them done. This technical guide is supported by a step-by-step computational tutorial accessible under https://tritexassembly.bitbucket.io/ . The TRITEX source code is hosted under this URL: https://bitbucket.org/tritexassembly .

An autoactive NB-LRR gene causes Rht13 dwarfism in wheat.

Semidwarfing genes have greatly increased wheat yields globally, yet the widely used gibberellin (GA)-insensitive genes Rht-B1b and Rht-D1b have disadvantages for seedling emergence. Use of the GA-sensitive semidwarfing gene Rht13 avoids this pleiotropic effect. Here, we show that Rht13 encodes a nucleotide-binding site/leucine-rich repeat (NB-LRR) gene. A point mutation in the semidwarf Rht-B13b allele autoactivates the NB-LRR gene and causes a height reduction comparable with Rht-B1b and Rht-D1b in diverse genetic backgrounds. The autoactive Rht-B13b allele leads to transcriptional up-regulation of pathogenesis-related genes including class III peroxidases associated with cell wall remodeling. Rht13 represents a new class of reduced height (Rht) gene, unlike other Rht genes, which encode components of the GA signaling or metabolic pathways. This discovery opens avenues to use autoactive NB-LRR genes as semidwarfing genes in a range of crop species, and to apply Rht13 in wheat breeding programs using a perfect genetic marker.

Discovery of stripe rust resistance with incomplete dominance in wild emmer wheat using bulked segregant analysis sequencing.

Durable crop disease resistance is an essential component of global food security. Continuous pathogen evolution leads to a breakdown of resistance and there is a pressing need to characterize new resistance genes for use in plant breeding. Here we identified an accession of wild emmer wheat (Triticum turgidum ssp. dicoccoides), PI 487260, that is highly resistant to multiple stripe rust isolates. Genetic analysis revealed resistance was conferred by a single, incompletely dominant gene designated as Yr84. Through bulked segregant analysis sequencing (BSA-Seq) we identified a 52.7 Mb resistance-associated interval on chromosome 1BS. Detected variants were used to design genetic markers for recombinant screening, further refining the interval of Yr84 to a 2.3-3.3 Mb in tetraploid wheat genomes. This interval contains 34 candidate genes encoding for protein domains involved in disease resistance responses. Furthermore, KASP markers closely-linked to Yr84 were developed to facilitate marker-assisted selection for rust resistance breeding.

Main effect and epistatic QTL affecting spike shattering and association with plant height revealed in two spring wheat (Triticum aestivum L.) populations.

KEY MESSAGE: A major QTL on chromosome arm 4BS was associated with reduced spike shattering and reduced plant height in coupling phase, and a second major QTL associated with reduced spike shattering was detected on chromosome arm 5AL in the same wheat variety Carberry. Spike shattering can cause severe grain yield loss in wheat. Development of cultivars with reduced shattering but having easy mechanical threshability is the target of wheat breeding programs. This study was conducted to determine quantitative trait loci (QTL) associated with shattering resistance, and epistasis among QTL in the populations Carberry/AC Cadillac and Carberry/Thatcher. Response of the populations to spike shattering was evaluated near Swift Current, SK, in four to five environments. Plant height data recorded in different locations and years were used to determine the relationship of the trait with spike shattering. Each population was genotyped and mapped with the wheat 90 K Illumina iSelect SNP array. Main effect QTL were analyzed by MapQTL 6, and epistatic interactions between main effect QTL were determined by QTLNetwork 2.0. Correlations between height and shattering ranged from 0.15 to 0.49. Carberry contributed two major QTL associated with spike shattering on chromosome arms 4BS and 5AL, detected in both populations. Carberry also contributed two minor QTL on 7AS and 7AL. AC Cadillac contributed five minor QTL on 1AL, 2DL, 3AL, 3DL and 7DS. Nine epistatic QTL interactions were identified, out of which the most consistent and synergistic interaction, that reduced the expression of shattering, occurred between 4BS and 5AL QTL. The 4BS QTL was consistently associated with reduced shattering and reduced plant height in the coupling phase. The present findings shed light on the inheritance of shattering resistance and provide genetic markers for manipulating the trait to develop wheat cultivars.

Tandem Protein Kinases Emerge as New Regulators of Plant Immunity.

Plant-pathogen interactions result in disease development in a susceptible host. Plants actively resist pathogens via a complex immune system comprising both surface-localized receptors that sense the extracellular space as well as intracellular receptors recognizing pathogen effectors. To date, the majority of cloned resistance genes encode intracellular nucleotide-binding leucine-rich repeat receptor proteins. Recent discoveries have revealed tandem kinase proteins (TKPs) as another important family of intracellular proteins involved in plant immune responses. Five TKP genes-barley Rpg1 and wheat WTK1 (Yr15), WTK2 (Sr60), WTK3 (Pm24), and WTK4-protect against devastating fungal diseases. Moreover, a large diversity and numerous putative TKPs exist across the plant kingdom. This review explores our current knowledge of TKPs and serves as a basis for future studies that aim to develop and exploit a deeper understanding of innate plant immunity receptor proteins.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Rhizosphere shotgun metagenomic analyses fail to show differences between ancestral and modern wheat genotypes grown under low fertilizer inputs.

It is thought that modern wheat genotypes have lost their capacity to associate with soil microbes that would help them acquire nutrients from the soil. To test this hypothesis, ten ancestral and modern wheat genotypes were seeded in a field experiment under low fertilization conditions. The rhizosphere soil was collected, its DNA extracted and submitted to shotgun metagenomic sequencing. In contrast to our hypothesis, there was no significant difference in the global rhizosphere metagenomes of the different genotypes, and this held true when focusing the analyses on specific taxonomic or functional categories of genes. Some genes were significantly more abundant in the rhizosphere of one genotype or another, but they comprised only a small portion of the total genes identified and did not affect the global rhizosphere metagenomes. Our study shows for the first time that the rhizosphere metagenome of wheat is stable across a wide variety of genotypes when growing under nutrient poor conditions.

The effects of crop attributes, selection, and recombination on Canadian bread wheat molecular variation.

Cultivated germplasm provides an opportunity to investigate how crop agronomic traits, selection for major genes, and differences in crossing-over rates drive patterns of allelic variation. To identify how these factors correlated with allelic variation within a collection of cultivated bread wheat (Triticum aestivum L.), we generated genotypes for 388 accessions grown in Canada over the past 170 yr using filtered single nucleotide polymorphism (SNP) calls from an Illumina Wheat iSelect 90K SNP-array. Entries' breeding program, era of release, grain texture, kernel color, and growth habit contributed to allelic differentiation. Allelic diversity and linkage disequilibrium (LD) of markers flanking some major loci known to affect traits such as gluten strength, growth habit, and grain color were consistent with selective sweeps. Nonetheless, some flanking markers of major loci had low LD and high allelic diversity. Positive selection may have acted upon homoeologous genes that had significant enrichment for the gene ontology terms 'response-to-auxin' and 'response-to-wounding.' Long regions of LD, spanning approximately one-third the length of entire chromosomes, were associated with many pericentromeric regions. These regions were also characterized by low diversity. Enhancing recombination across these regions could generate novel allele combinations to accelerate Canadian wheat improvement.

Long-read sequence assembly: a technical evaluation in barley.

Sequence assembly of large and repeat-rich plant genomes has been challenging, requiring substantial computational resources and often several complementary sequence assembly and genome mapping approaches. The recent development of fast and accurate long-read sequencing by circular consensus sequencing (CCS) on the PacBio platform may greatly increase the scope of plant pan-genome projects. Here, we compare current long-read sequencing platforms regarding their ability to rapidly generate contiguous sequence assemblies in pan-genome studies of barley (Hordeum vulgare). Most long-read assemblies are clearly superior to the current barley reference sequence based on short-reads. Assemblies derived from accurate long reads excel in most metrics, but the CCS approach was the most cost-effective strategy for assembling tens of barley genomes. A downsampling analysis indicated that 20-fold CCS coverage can yield very good sequence assemblies, while even five-fold CCS data may capture the complete sequence of most genes. We present an updated reference genome assembly for barley with near-complete representation of the repeat-rich intergenic space. Long-read assembly can underpin the construction of accurate and complete sequences of multiple genomes of a species to build pan-genome infrastructures in Triticeae crops and their wild relatives.

Chromosome-scale genome assembly provides insights into rye biology, evolution and agronomic potential.

Rye (Secale cereale L.) is an exceptionally climate-resilient cereal crop, used extensively to produce improved wheat varieties via introgressive hybridization and possessing the entire repertoire of genes necessary to enable hybrid breeding. Rye is allogamous and only recently domesticated, thus giving cultivated ryes access to a diverse and exploitable wild gene pool. To further enhance the agronomic potential of rye, we produced a chromosome-scale annotated assembly of the 7.9-gigabase rye genome and extensively validated its quality by using a suite of molecular genetic resources. We demonstrate applications of this resource with a broad range of investigations. We present findings on cultivated rye's incomplete genetic isolation from wild relatives, mechanisms of genome structural evolution, pathogen resistance, low-temperature tolerance, fertility control systems for hybrid breeding and the yield benefits of rye-wheat introgressions.

Advanced domestication: harnessing the precision of gene editing in crop breeding.

Human population growth has increased the demand for food crops, animal feed, biofuel and biomaterials, all the while climate change is impacting environmental growth conditions. There is an urgent need to develop crop varieties which tolerate adverse growth conditions while requiring fewer inputs. Plant breeding is critical to global food security and, while it has benefited from modern technologies, it remains constrained by a lack of valuable genetic diversity, linkage drag, and an effective way to combine multiple favourable alleles for complex traits. CRISPR/Cas technology has transformed genome editing across biological systems and promises to transform agriculture with its high precision, ease of design, multiplexing ability and low cost. We discuss the integration of CRISPR/Cas-based gene editing into crop breeding to advance domestication and refine inbred crop varieties for various applications and growth environments. We highlight the use of CRISPR/Cas-based gene editing to fix desirable allelic variants, generate novel alleles, break deleterious genetic linkages, support pre-breeding and for introgression of favourable loci into elite lines.

A Combination of Leaf Rust Resistance Genes, Including Lr34 and Lr46, Is the Key to the Durable Resistance of the Canadian Wheat Cultivar, Carberry.

The hexaploid spring wheat cultivar, Carberry, was registered in Canada in 2009, and has since been grown over an extensive area on the Canadian Prairies. Carberry has maintained a very high level of leaf rust (Puccinia triticina Eriks.) resistance since its release. To understand the genetic basis of Carberry's leaf rust resistance, Carberry was crossed with the susceptible cultivar, Thatcher, and a doubled haploid (DH) population of 297 lines was generated. The DH population was evaluated for leaf rust in seven field environments at the adult plant stage. Seedling and adult plant resistance (APR) to multiple virulence phenotypes of P. triticina was evaluated on the parents and the progeny population in controlled greenhouse studies. The population was genotyped with the wheat 90 K iSelect single nucleotide polymorphism (SNP) array, and quantitative trait loci (QTL) analysis was performed. The analysis using field leaf rust response indicated that Carberry contributed nine QTL located on chromosomes 1B, 2B (2 loci), 2D, 4A, 4B, 5A, 5B, and 7D. The QTL located on 1B, 2B, 5B, and 7D chromosomes were observed in two or more environments, whereas the remainder were detected in single environments. The resistance on 1B, detected in five environments, was attributed to Lr46 and on 7D, detected in seven environments to Lr34. The first 2B QTL corresponded with the adult plant gene, Lr13, while the second QTL corresponded with Lr16. The seedling analysis showed that Carberry carries Lr2a, Lr16, and Lr23. Five epistatic effects were identified in the population, with synergistic interactions being observed for Lr34 with Lr46, Lr16, and Lr2a. The durable rust resistance of Carberry is attributed to Lr34 and Lr46 in combination with these other resistance genes, because the resistance has remained effective even though the P. triticina population has evolved virulent to Lr2a, Lr13, Lr16, and Lr23.