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Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat (Triticum aestivum L.) worldwide. Xiaoyan 78829, a partial amphidiploid developed by crossing common wheat with Thinopyrum intermedium, is immune to wheat stripe rust. To transfer the resistance gene of this excellent germplasm resource to wheat, the translocation line WTT11 was produced by pollen irradiation and assessed for immunity to stripe rust races CYR32, CYR33 and CYR34. A novel stripe rust-resistance locus derived from Th. intermedium was confirmed by linkage and diagnostic marker analyses. Molecular cytogenetic analyses revealed that WTT11 carries a TTh·2DL translocation. The breakpoint of 1B was located at 95.5 MB, and the alien segments were found to be homoeologous to wheat-group chromosomes 6 and 7 according to a wheat660K single-nucleotide polymorphism (SNP) array analysis. Ten previously developed PCR-based markers were confirmed to rapidly trace the alien segments of WTT11, and 20 kompetitive allele-specific PCR (KASP) markers were developed to enable genotyping of Th. intermedium and common wheat. Evaluation of agronomic traits in two consecutive crop seasons uncovered some favorable agronomic traits in WTT11, such as lower plant height and longer main panicles, that may be applicable to wheat improvement. As a novel genetic resource, the new resistance locus may be useful for wheat disease-resistance breeding.

Premise Nucleic acid integrity can be compromised under many abiotic stresses. To date, however, few studies have considered whether nucleic acid damage and damage repair play a role in cold-stress adaptation. A further insufficiently explored question concerns how age affects cold stress adaptation among mature perennials. As a plant ages, the optimal trade-off between growth and stress tolerance may shift. Methods Oxidative damage to RNA and expression of genes involved in DNA repair were compared in multiple mature cohorts of Thinopyrum intermedium (an emerging perennial cereal) and in wheat and barley under intermittent freezing stress and under nonfreezing conditions. Activity of glutathione peroxidase (GPX) and four other antioxidative enzymes was also measured under these conditions. DNA repair genes included photolyases involved in repairing ultraviolet-induced damage and two genes involved in repairing oxidatively induced damage (ERCC1, RAD23). Results Freezing stress was accompanied by large increases in photolyase expression and ERCC1 expression (in wheat and Thinopyrum) and in GPX and GR activity (particularly in Thinopyrum). This is the first report of DNA photolyases being overexpressed under freezing stress. Older Thinopyrum had lower photolyase expression and less freezing-induced overexpression of ERCC1. Younger Thinopyrum plants sustained more oxidative damage to RNA. Conclusions Overexpression of DNA repair genes is an important aspect of cold acclimation. When comparing adult cohorts, aging was associated with changes in the freezing stress response, but not with overall increases or decreases in stress tolerance.

Thinopyrum intermedium (2n = 6x = 42, E1E1E2E2XX) serves as an important gene source of desirable traits for genetic improvement of wheat cultivars resistant to stresses. This study used the comparative proteomic approach to identify stress defense related proteins in the developing grains of common wheat (Zhongmai 8601)-Thinopyron intermedium 7XL/7DS translocation line YW642 and to explore their potential values for improving wheat stress resistance. Two-dimensional electrophoresis identified 124 differentially accumulated protein spots representing 100 unique proteins, which mainly participated in stress defense, energy metabolism, protein metabolism and folding and storage protein synthesis. Among these, 16 were unique and 35 were upregulated in YW642. The upregulated DAPs were mainly involved in biotic and abiotic stress defense. Further cis-elements analysis of these stress-related DAP genes revealed that phytohormone responsive elements such as ABREs, G-box, CGTCA-motif and TGACG-motif, and environment responsive element As-1 were particularly abundant, which could play important roles in response to various stressors. Transcription expression analysis by RNA-seq and qRT-PCR demonstrated a large part of the stress-related DAP genes showed an upregulated expression in the early-to-middle stages of grain development. Our results proved that Thinopyron intermedium contains abundant stress responsive proteins that have potential values for the genetic improvement of wheat stress resistance.

Key Message Paternity assignment and genome-wide association analyses for fertility were applied to aThinopyrum intermediumbreeding program. A lack of progeny between combinations of parents was associated with loci near self-incompatibility genes. Abstract In outcrossing species such as intermediate wheatgrass (IWG, Thinopyrum intermedium), polycrossing is often used to generate novel recombinants through each cycle of selection, but it cannot track pollen-parent pedigrees and it is unknown how self-incompatibility (SI) genes may limit the number of unique crosses obtained. This study investigated the potential of using next-generation sequencing to assign paternity and identify putative SI loci in IWG. Using a reference population of 380 individuals made from controlled crosses of 64 parents, paternity was assigned with 92% agreement using Cervus software. Using this approach, 80% of 4158 progeny (n = 3342) from a polycross of 89 parents were assigned paternity. Of the 89 pollen parents, 82 (92%) were represented with 1633 unique full-sib families representing 42% of all potential crosses. The number of progeny per successful pollen parent ranged from 1 to 123, with number of inflorescences per pollen parent significantly correlated to the number of progeny (r = 0.54, p

Stripe rust is an important disease in wheat, and development of genetic resistance in cultivars is an effective approach to control the disease. Wild species of wheat, such as Thinopyrum intermedium, are an excellent gene source for wheat improvement. In this study, two stripe rust-resistant wheat–Th. intermedium chromosome translocation lines, CH4131 and CH4132, were characterized by cytogenetic and pathological methods. The introgressed chromosome fragment was tagged using amplified fragment-length polymorphism-derived sequence-characterized amplified region (SCAR) markers and intron targeting markers, indicating that CH4131 and CH4132 both possess a homologous group 3 chromatin of Th. intermedium. Genomic in situ hybridization results suggested that a very small Th. intermedium chromosome segment was translocated to the terminal region of wheat 1BS for both lines, forming a configuration of T3Ai-1BS.1BL. The two translocation lines were resistant to stripe rust, and the resistance gene, temporarily designated YrCH-1BS, was likely derived from Th. intermedium. The translocated chromosome fragments have no genetic linkage drag to agronomic performance. The grain quality indexes of these two translocations were higher than local wheat varieties. Therefore, CH4131 and CH4132 could be used as potential gene sources in wheat improvement programs. The SCAR markers are useful to select stripe rust resistance from Th. intermedium.

Thinopyrum intermedium (2n = 6x = 42, JJJsJsStSt) has been hybridized extensively with common wheat and has proven to be a valuable germplasm source for improving disease resistance, quality attributes, and yield potential in wheat. We characterized new disease resistant wheat-Th. intermedium derivatives A1082 and A5-5 using sequential multi-color fluorescence in situ hybridization (mc-FISH), genomic in situ hybridization (GISH), PCR-based landmark unique gene (PLUG) and intron targeting (IT) markers. A1082 was identified as a wheat-Th. intermedium 3J disomic addition line, and A5-5 was a T4BS·5JsL homozygous Robertsonian translocation line. Seventy-one and 106 pairs of primers amplified Th. intermedium-specific bands allowing chromosomes 3J and 5Js to be tracked, respectively. A new oligonucleotide probe, Oligo-6H-2-100, was developed for FISH labeling of the subterminal region of the long arm of chromosome 5Js. Both lines were highly resistant to stripe rust pathogen races prevalent in Chinese field screening nurseries. A5-5 also displayed a significant increase in tiller number compared to its wheat parent. The new lines can be exploited as useful germplasms for wheat improvement. Keywords: Disease resistance, FISH, GISH, Stripe rust, Triticum aestivum

Powdery mildew caused by the fungal pathogen Blumeria graminis f. sp. tritici (Bgt) is an econo-mically important disease of common wheat T. aestivum L. One of the most effective and environmentally important ways of protection of wheat against Bgt is cultivation of the varieties with genetic resistance. The aim of this work was to study the genetic diversity of Russian spring wheat varieties for the powdery mildew resistance loci. Phytopathological evaluation of 97 wheat varieties showed that no more than 10% of the varieties have low level of susceptibility to the Bgt population specific to the Western Siberian region. Association mapping carried out on the basis of SNP genotyping and phytopathological testing during three environmental seasons identified eight loci in chromosomes 1AL, 1DS, 2BL, 5AS, 5DS, 6AL, 6DL, and 7AL. A high impact to the phenotypic manifestation of the trait was established for genetic factors localized in chromosomes 5AS, 6AL, and 6DL. The long arm of chromosome 6D contains the gene Pm6Ai=2, which was introduced from wheatgrass Thinopyrum intermedium and provides effective protection against the powdery mildew pathogen. On the basis of comparative analysis of the chromosomal localization of the known Pm resistance genes and loci mapped in this work, it was assumed that QTLs in chromosomes 1DS, 5AS, and 6AL are novel, not previously described resistance loci. The obtained results can be used in breeding programs for selection of target loci and for development of molecular markers specific to Bgt resistance loci.

Barley yellow dwarf (BYD) is a major virus disease which dramatically reduces wheat yield. Introducing BYD resistance genes into commercial varieties has been proven to be effective in reducing damage caused by barley yellow dwarf virus (BYDV). However, only one major resistance gene is readily deployable for breeding; Bdv2 derived from Thinopyrum intermedium is deployed as a chromosomal translocation. In this study, a double haploid (DH) population was developed from a cross between XuBYDV (introduced from China showing very good resistance to BYD) and H-120 (a BYD-sensitive Chinese accession), and was used to identify QTL for BYD resistance. The population was genotyped using an Infinium iSelect bead chip array targeting 90K gene-based SNPs. The disease resistance of DH lines inoculated with BYDV was assessed at the heading stage. The infections were assessed by tissue blot immunoassay (TBIA). Three new QTL were identified on chromosomes 5A, 6A, and 7A for both symptom and TBIA, with all three resistance alleles being inherited from XuBYDV. Some DH lines with the resistance alleles from all three QTL showed high level resistance to BYD. These new QTL will be useful in breeding programs for pyramiding BYD resistance genes.

Aim. Thinopyrum intermedium , a wild wheat relative, for some reasons is on interest for its involvement to the introgressive hybridization with wheat. The basis for the purposeful work in this direction should be the information about the biological and genetic properties of the species, including the mating system. The mating system can be identified based on the genotyping of plants in populations for a polymorphic gene with codominant inheritance. Gene Glu was used as such a codominant gene. Methods. Cytological (determination of the chromosome number in metaphases of mitosis), electrophoresis of glutenin storage proteins, comparative analysis of electrophoretic spectra, and the population genetic methods for calculating the frequencies of genes and genotypes . Results. Plants were hexaploid, with 2n = 42 chromosomes. In a sample of fifty plants, a genetic control of components of the glutenin spectra has been established. They are controlled by three independent genes: Glu-Thi1-2 with four alleles 1-2 (cluster), 1, 2, 0, Glu-Thi1-4 withfour alleles 1-4 (cluster), 1, 4, 0, and the three-allele gene Glu-Thi1-3-5 . The participation of the three genes in the formation of the electrophoretic spectrum of glutenins is consistent with the hexaploid nature of Thinopyrum intermedium , which follows from the established number of 42 chromosomes. The information about the genetic control of glutenins is the basis for determining the mating system of this species. A simplified three-allele model of genetic control of glutenins’ electrophoretic spectra was used for determination of the mating system of Thinopyrum intermedium. Conclusions. Plants of the studied Thinopyrum intermedium population have a mixed mating system and the probability of self-pollination ranging from 0,50 to 0,81, depending on the Glu gene, which is used for determination of heterozygosity of the plants. This biological characteristic is positive for prospective use of this species for wide hybridization.

Intermediate wheatgrass (Thinopyrum intermedium, IWG) is a perennial grain crop with high biomass and grain yield, long seeds, and resistance to pests and diseases. It also reduces soil erosion, nitrate and mineral leaching into underground water tables, and sequesters carbon in its roots. The domestication timeline of IWG as a grain crop spans only 3 decades, hence it lags annual grain crops in yield and seed characteristics. One approach to improve its agronomic traits is by using molecular markers to uncover marker-trait associations. In this study, we performed association mapping on IWG breeding germplasm from the third recurrent selection cycle at the University of Minnesota. The IWG population was phenotyped in St Paul, MN in 2017 and 2018, and in Crookston, MN in 2018 for grain yield, seed length, width and weight, spike length and weight, and number of spikelets per spike. Strong positive correlations were observed among most trait pairs, with correlations as high as 0.76. Genotyping using high throughput sequencing identified 8,899 high-quality genome-wide SNPs which were combined with phenotypic data in association mapping to discover regions associated with the yield component traits. We detected 154 genetic loci associated with these traits of which 19 were shared between at least two traits. Prediction of breeding values using significant loci as fixed effects in genomic selection model improved predictive abilities by up to 14%. Genetic mapping of agronomic traits followed by using genomic selection to predict breeding values can assist breeders in selecting superior genotypes to accelerate IWG domestication.

Background Interspecific hybridisation resulting in polyploidy is one of the major driving forces in plant evolution. Here, we present data from the molecular cytogenetic analysis of three cytotypes of Elytrigia ×mucronata using sequential fluorescence (5S rDNA, 18S rDNA and pSc119.2 probes) and genomic in situ hybridisation (four genomic probes of diploid taxa, i.e., Aegilops, Dasypyrum, Hordeum and Pseudoroegneria). Results The concurrent presence of Hordeum (descended from E. repens) and Dasypyrum + Aegilops (descended from E. intermedia) chromosome sets in all cytotypes of E. ×mucronata confirmed the assumed hybrid origin of the analysed plants. The following different genomic constitutions were observed for E. ×mucronata. Hexaploid plants exhibited three chromosome sets from Pseudoroegneria and one chromosome set each from Aegilops, Hordeum and Dasypyrum. Heptaploid plants harboured the six chromosome sets of the hexaploid plants and an additional Pseudoroegneria chromosome set. Nonaploid cytotypes differed in their genomic constitutions, reflecting different origins through the fusion of reduced and unreduced gametes. The hybridisation patterns of repetitive sequences (5S rDNA, 18S rDNA, and pSc119.2) in E. ×mucronata varied between and within cytotypes. Chromosome alterations that were not identified in the parental species were found in both heptaploid and some nonaploid plants. Conclusions The results confirmed that both homoploid hybridisation and heteroploid hybridisation that lead to the coexistence of four different haplomes within single hybrid genomes occur in Elytrigia allopolyploids. The chromosomal alterations observed in both heptaploid and some nonaploid plants indicated that genome restructuring occurs during and/or after the hybrids arose. Moreover, a specific chromosomal translocation detected in one of the nonaploids indicated that it was not a primary hybrid. Therefore, at least some of the hybrids are fertile. Hybridisation in Triticeae allopolyploids clearly and significantly contributes to genomic diversity. Different combinations of parental haplomes coupled with chromosomal alterations may result in the establishment of unique lineages, thus providing raw material for selection. Electronic supplementary material The online version of this article (10.1186/s12870-019-1806-y) contains supplementary material, which is available to authorized users.

Thinopyrum intermedium and barley are two close relatives of wheat and carry many genes that are potentially valuable for the improvement of various wheat traits. In this study we created wheat double substitution lines by hybridizing different wheat–Th. intermedium and wheat–barley disomic alien substitution lines, with the aim of using genes in Th. intermedium and barley for wheat breeding and investigating the genetic behavior of alien chromosomes and their wheat homoeologs. As expected, we obtained two types of wheat double substitution lines, 2D2Ai#2(2B)2H(2A) and 2A2Ai#2(2B)2H(2D), in which different group 2 wheat chromosomes were replaced by barley chromosome 2H and Th. intermedium chromosome 2Ai#2. The new materials were characterized using molecular markers, genomic in situ hybridization (GISH), and fluorescent in situ hybridization (FISH). GISH and FISH experiments revealed that the double substitution lines harbor 42 chromosomes including 38 wheat chromosomes, a pair of barley chromosomes, and a pair of Th. intermedium chromosomes. Analysis using specific DNA markers showed that two pairs of wheat homoeologous group 2 chromosomes in the new lines were substituted by a pair of 2H and a pair of 2Ai#2 chromosomes. Chromosome 2H showed a higher transmission rate than 2Ai#2, and both chromosomes were preferentially transmitted between generations via female gametes. Evaluation of botanic and agronomic traits demonstrated that, compared with their parents, the new lines showed similar growth habits and plant type but differences in plant height, flowering date, and self-fertility. Cytological observations using different probes suggested that the double substitution lines showed nearly normal genetic behavior before and during meiosis. The novel substitution lines can potentially be used in wheat meiosis research and breeding programs. Keywords: Wheat, Double substitution lines, Molecular markers, Transmission rate of alien chromosomes, Genomic in situ hybridization

In an era of constrained and depleted natural resources, perennial grains could provide sustainable food production along with beneficial ecosystem services like reduced erosion and increased atmospheric carbon capture. Intermediate wheatgrass (IWG) [Thinopyrum intermedium (Host) Barkworth & D. R. Dewey subsp. intermedium] has been undergoing continuous breeding for domestication to develop a perennial grain crop since the 1980s. As a perennial, IWG has required 2–5 yr per selection generation, but starting in 2017, genomic selection (GS) was initiated in the breeding program at The Land Institute, Salina, KS (TLI), enabling one complete cycle per year. For each cycle, ∼4,000 seedlings were profiled using genotyping‐by‐sequencing (GBS) and genomic estimated breeding values (GEBVs) were calculated. Selection based on GEBVs identified ∼100 individuals to advance as parents each generation, while validation populations of 1,000–1,200 genets for GS model training were also selected using the genomic relationship matrix to represent genetic diversity in each cycle. The selected parents were randomly intermated in a greenhouse crossing block to form the subsequent cycle, while the validation populations were transplanted to irrigated and nonirrigated field sites for phenotypic evaluations in the following years. For priority breeding traits of seed mass, free threshing, and nonshattering, correlations between predicted values and observed data were >.5. The realized selection differential ranged from 11–23% for selected traits, and the expected genetic gains for these traits, including spike yield, ranged from 6 to 14% per year. Genomic selection is a powerful tool to speed the domestication and development of IWG and other perennial crops with extended breeding timelines.

Thinopyrum intermedium (2n = 6x = 42, JJJSJSStSt) is one of the important resources for the wheat improvement. So far, a few Th. intermedium (Thi)-specific molecular markers have been reported, but the number is far from enough to meet the need of identifying alien fragments in wheat-Th. intermedium hybrids. In this study, 5,877,409 contigs were assembled using the Th. intermedium genotyping-by-sequencing (GBS) data. We obtained 5,452 non-redundant contigs containing mapped Thi-GBS markers with less than 20% similarity to the wheat genome and developed 2,019 sequence-tagged site (STS) molecular markers. Among the markers designed, 745 Thi-specific markers with amplification products in Th. intermedium but not in eight wheat landraces were further selected. The distribution of these markers in different homologous groups of Th. intermedium varied from 47 (7/12/28 on 6J/6St/6JS) to 183 (54/62/67 on 7J/7St/7JS). Furthermore, the effectiveness of these Thi-specific markers was verified using wheat-Th. intermedium partial amphidiploids, addition lines, substitution lines, and translocation lines. Markers developed in this study provide a convenient, rapid, reliable, and economical method for identifying Th. intermedium chromosomes in wheat. In addition, this set of Thi-specific markers can also be used to estimate genetic and physical locations of Th. intermedium chromatin in the introgression lines, thus providing valuable information for follow-up studies such as alien gene mining.

Owing to the excellent resistance to abiotic and biotic stress, Thionpyrum intermedium (2n = 6x = 42, JJJsJsStSt) and Thinopyrum ponticum (2n = 10x = 70) are both widely utilized in wheat germplasm innovation programs. Disomic substitution lines (DSLs) carrying one pair of alien chromosomes are valuable bridge materials for novel genes transmission. In this study, six wheat-Thinopyrum DSLs were derived from crosses between Abbondanza nullisomic lines (2n = 40) and two octoploid Trititrigia lines (2n = 8x = 56), characterized by a sequential fluorescence in situ hybridization (FISH)-genome in situ hybridization (GISH), a multicolor GISH (mc-GISH), and an analysis of wheat 15K SNP array combined with molecular marker selection. ES-9 and ES-10 were two wheat- Th. ponticum disomic substitution lines, DS2St (2A) and DS3St (3D). While ES-23, ES-24, ES-25, and ES-26 were four wheat- Th. intermedium disomic substitution lines, DS2St (2A), DS3St (3D), DS2St (2B), DS2St (2D). The FISH karyotypes of Th. ponticum 2St/3St chromosomes were well coincident with the ones of Th. intermedium. The chromosome configurations of F1 hybrids derived from crosses between ES-23 and ES-9, as well as ES-24 and ES-10 were mostly formed 21Ⅱ. Four St-chromosome-specific markers were developed by specific-locus amplified fragment sequencing (SLAF-seq). Additionally, the substitution lines containing chromosome 2St conferred higher thousand-kernel weight and stripe rust resistance at adult stages, while the substitution lines containing chromosome 3St were highly resistant to stripe rust at all stages. Therefore, these six substitution lines could serve as useful bridging parents for wheat genetic improvement.

Perennial grain crops have the potential to improve agricultural sustainability but few existing species produce sufficient grain yield to be economically viable. The outcrossing, allohexaploid, and perennial forage species intermediate wheatgrass (IWG) [Thinopyrum intermedium (Host) Barkworth & D. R. Dewey] has shown promise in undergoing direct domestication as a perennial grain crop using phenotypic and genomic selection. However, decades of selection will be required to achieve yields on par with annual small‐grain crops. Marker‐aided selection could accelerate progress if important genomic regions associated with domestication were identified. Here we use the IWG nested association mapping (NAM) population, with 1,168 F1 progeny across 10 families to dissect the genetic control of brittle rachis, floret shattering, and threshability. We used a genome‐wide association study (GWAS) with 8,003 single nucleotide polymorphism (SNP) markers and linkage mapping—both within‐family and combined across families—with a robust phenotypic dataset collected from four unique year‐by‐location combinations. A total of 29 quantitative trait loci (QTL) using GWAS and 20 using the combined linkage analysis were detected, and most large‐effect QTL were in common across the two analysis methods. We reveal that the genetic control of these traits in IWG is complex, with significant QTL across multiple chromosomes, sometimes within and across homoeologous groups and effects that vary depending on the family. In some cases, these QTL align within 216 bp to 31 Mbp of BLAST hits for known domestication genes in related species and may serve as precise targets of selection and directions for further study to advance the domestication of IWG.

Background Thinopyrum intermedium (2n = 6x = 42) is an important wild perennial Triticeae species exhibiting many potentially favorable traits for wheat improvement. Wheat-Th. intermedium partial amphiploids serve as a bridge to transfer desirable genes from Th. intermedium into common wheat. Results Three octoploid Trititrigia accessions (TE261–1, TE266–1, and TE346–1) with good resistances to stripe rust, powdery mildew and aphids were selected from hybrid progenies between Th. intermedium and the common wheat variety ‘Yannong 15’ (YN15). Genomic in situ hybridization (GISH), fluorescence in situ hybridization (FISH) and multicolor GISH (McGISH) analyses demonstrated that the three octoploid Trititrigia possess 42 wheat chromosomes and 14 Th. intermedium chromosomes. The 14 alien (Th. intermedium) chromosomes belong to a mixed genome consisting of J-, JS- and St-genome chromosomes rather than a single J, JS or St genome. Different types of chromosomal structural variation were also detected in the 1A, 6A, 6B, 2D and 7D chromosomes via FISH, McGISH and molecular marker analysis. The identity of the alien chromosomes and the variationes in the wheat chromosomes in the three Trititrigia octoploids were also different. Conclusions The wheat-Th. intermedium partial amphiploids possess 14 alien chromosomes which belong to a mixed genome consisting of J-, JS- and St- chromosomes, and 42 wheat chromosomes with different structural variations. These accessions could be used as genetic resources in wheat breeding for the transfer of disease and pest resistance genes from Th. intermedium to common wheat.

Intermediate wheatgrass (Thinopyrum intermedium) is an outcrossing, cool season grass species currently undergoing direct domestication as a perennial grain crop. Though many traits are selection targets, understanding the genetic architecture of those important for local adaptation may accelerate the domestication process. Nested association mapping (NAM) has proven useful in dissecting the genetic control of agronomic traits many crop species, but its utility in primarily outcrossing, perennial species has yet to be demonstrated. Here, we introduce an intermediate wheatgrass NAM population developed by crossing ten phenotypically divergent donor parents to an adapted common parent in a reciprocal manner, yielding 1,168 F1 progeny from 10 families. Using genotyping by sequencing, we identified 8,003 SNP markers and developed a population-specific consensus genetic map with 3,144 markers across 21 linkage groups. Using both genomewide association mapping and linkage mapping combined across and within families, we characterized the genetic control of flowering time. In the analysis of two measures of maturity across four separate environments, we detected as many as 75 significant QTL, many of which correspond to the same regions in both analysis methods across 11 chromosomes. The results demonstrate a complex genetic control that is variable across years, locations, traits, and within families. The methods were effective at detecting previously identified QTL, as well as new QTL that align closely to the well-characterized flowering time orthologs from barley, including Ppd-H1 and Constans. Our results demonstrate the utility of the NAM population for understanding the genetic control of flowering time and its potential for application to other traits of interest.

A physical map of Thinopyrum intermedium chromosome 7J was constructed using translocation mapping, and a new seedling purple coleoptile gene was mapped to the bin of FL 0.35–0.63 of 7JS. Thinopyrum intermedium (2n = 6x = 42, JJJsJsStSt), a wild relative of common wheat, harbors numerous beneficial genes for wheat improvement. Previous studies showed that wheat–Th. intermedium partial amphiploid TAF46 and its derived addition line L1 had a purple coleoptile, which was derived from Th. intermedium chromosome 7J. To identify and physically map the purple coleoptile gene, 12 wheat–Th. intermedium 7J translocation lines were analyzed by sequential multicolor fluorescence in situ hybridization (mc-FISH), PCR-based landmark unique gene (PLUG) and intron targeting (IT) markers. A physical map of the 7J chromosome was constructed, consisting of eight chromosomal bins with 89 markers. Seedling evaluation of the coleoptile colors of all tested materials indicated that the purple coleoptile gene was located to the bin with a fraction length (FL) of 0.35–0.63 on chromosome 7JS. Furthermore, based on the syntenic relationships between Th. intermedium and wheat chromosomes, we developed a new chromosome 7J-specific EST-PCR marker from the chromosomal region corresponding to the purple coleoptile gene through the Triticeae multi-omics database. The approach of designing chromosome-specific markers has facilitated fine mapping of the Thinopyrum-specific purple coleoptile gene, and these translocation lines will be valuable for studying the function of the purple coleoptile gene in anthocyanin biosynthesis.

The development of perennial grain crops is driven by the vision of simultaneous food production and enhanced ecosystem services. Typically, perennial crops like intermediate wheatgrass (IWG)[Thinopyrum intermedium (Host) Barkworth & D.R Dewey] have low seed yield and other detrimental traits. Next‐generation sequencing has made genomic selection (GS) a tractable and viable breeding method. To investigate how an IWG breeding program may use GS, we evaluated 3,658 genets over 2 yr for 46 traits to build a training population. Six statistical models were used to evaluate the non‐replicated data, and a model using autoregressive order 1 (AR1) spatial correction for rows and columns combined with the genomic relationship matrix provided the highest estimates of heritability. Genomic selection models were built from 18,357 single nucleotide polymorphism markers via genotyping‐by‐sequencing, and a 20‐fold cross‐validation showed high predictive ability for all traits (r > .80). Predictive abilities improved with increased training population size and marker numbers, even with larger amounts of missing data per marker. On the basis of these results, we propose a GS breeding method that is capable of completing one cycle per year compared with a minimum of 2 yr per cycle with phenotypic selection. We estimate that this breeding approach can increase the rate of genetic gain up to 2.6× above phenotypic selection for spike yield in IWG, allowing GS to enable rapid domestication and improvement of this crop. These breeding methods should be transferable to other species with similar long breeding cycles or limited capacity for replicated observations.

Starch, as the main component of grain in cereals, serves as the major source of calories in staple food and as a raw material for industry. As the technological and digestive properties of starch depend on its content, the management of its components, amylose and amylopectin, is of great importance. The starch properties of wheat grain can be attuned using allelic variations of genes, including granule-bound starch synthase I (GBSS I), or Wx. The tertiary gene pool, including wheatgrass (Thinopyrum) species, provides a wide spectrum of genes-orthologs that can be used to increase the allelic diversity of wheat genes by wide hybridization. Octaploid partial wheat&ndash, wheatgrass hybrids (amphidiploids, WWGHs) combine the complete genome of bread wheat (BBAADD), and a mixed genome from the chromosomes of intermediate wheatgrass (Thinopyrum intermedium, genomic composition JrJrJvsJvsStSt) and tall wheatgrass (Th. ponticum, JJJJJJJsJsJsJs). Thus, WWGHs may carry Wx genes not only of wheat (Wx-B1, Wx-A1 and Wx-D1) but also of wheatgrass origin. We aimed to assess the level of amylose in starch and investigate the polymorphism of Wx genes in 12 accessions of WWGHs. Additionally, we characterized orthologous Wx genes in the genomes of wild wheat-related species involved in the development of the studied WWGHs, Th. intermedium and Th. ponticum, as well as in the putative donors of their subgenomes, bessarabian wheatgrass (Th. bessarabicum, JbJb) and bluebunch wheatgrass (Pseudoroegneria stipifolia, St1St1St2St2). Although no significant differences in amylose content were found between different WWGH accessions, SDS-PAGE demonstrated that at least two WWGHs have an additional band. We sequenced the Wx gene-orthologs in Th. bessarabicum, P. stipifolia, Th. intermedium and Th. ponticum, and developed a WXTH marker that can discriminate the Thinopyrum Wx gene in the wheat background, and localized it to the 7E chromosome in Th. elongatum. Using the WXTH marker we revealed the allelic polymorphism of the Thinopyrum Wx gene in the studied WWGHs. The applicability of Thinopyrum Wx genes in wheat breeding and their effect on starch quality are discussed.

The cytogenetic study of wide hybrids of wheat has both practical and fundamental values. Partial wheat-wheatgrass hybrids (WWGHs) are interesting as a breeding bridge to confer valuable genes to wheat genome, as well as a model object that contains related genomes of Triticeae. The development of cytogenetic markers is a process that requires long and laborious fluorescence in situ hybridization (FISH) testing of various probes before a suitable probe is found. In this study, we aimed to find an approach that allows to facilitate this process. Based on the data sequencing of Thinopyrum ponticum, we selected six tandem repeat (TR) clusters using RepeatExplorer2 pipeline and designed primers for each of them. We estimated the found TRs&rsquo, abundance in the genomes of Triticum aestivum, Thinopyrum ponticum, Thinopyrum intermedium and four different WWGH accessions using real-time qPCR, and localized them on the chromosomes of the studied WWGHs using fluorescence in situ hybridization. As a result, we obtained three tandem repeat cytogenetic markers that specifically labeled wheatgrass chromosomes in the presence of bread wheat chromosomes. Moreover, we designed and tested primers for these repeats, and demonstrated that they can be used as qPCR markers for quick and cheap monitoring of the presence of certain chromosomes of wheatgrass in breeding programs.

Genomic selection (GS) based recurrent selection methods were developed to accelerate the domestication of intermediate wheatgrass [IWG, Thinopyrum intermedium (Host) Barkworth & D.R. Dewey]. A subset of the breeding population phenotyped at multiple environments is used to train GS models and then predict trait values of the breeding population. In this study, we implemented several GS models that investigated the use of additive and dominance effects and G×E interaction effects to understand how they affected trait predictions in intermediate wheatgrass. We evaluated 451 genotypes from the University of Minnesota IWG breeding program for nine agronomic and domestication traits at two Minnesota locations during 2017-2018. Genet-mean based heritabilities for these traits ranged from 0.34 to 0.77. Using four-fold cross validation, we observed the highest predictive abilities (correlation of 0.67) in models that considered G×E effects. When G×E effects were fitted in GS models, trait predictions improved by 18%, 15%, 20%, and 23% for yield, spike weight, spike length, and free threshing, respectively. Genomic selection models with dominance effects showed only modest increases of up to 3% and were trait-dependent. Cross-environment predictions were better for high heritability traits such as spike length, shatter resistance, free threshing, grain weight, and seed length than traits with low heritability and large environmental variance such as spike weight, grain yield, and seed width. Our results confirm that GS can accelerate IWG domestication by increasing genetic gain per breeding cycle and assist in selection of genotypes with promise of better performance in diverse environments.

Aim. Compare structures of Glu genes, extracted from Th. intermedium plants grown in different microgeographic conditions. Methods. PCR with DNA samples of Th. intermedium and primers to Glu-1Dx and Glu-1Dy gene regions, sequencing and comparative analysis of polymorphic amplicons and sequences from databases. Results. Central fragments of Glu-1Dx and Glu-1Dy from plants grown in different microgeographic conditions had significant differences compared to corresponding sequences of Aegilops tauschii (Glu-1Dx), Thinopyrum intermedium (Glu-1St*2x), Triticum aestivum (Glu-1Dy) obtained from databases. Both genes had point mutations, deletions and insertions, which transformed glutamine coding triplets characteristic for storage proteins to stop-codons, also codons for cysteine, methionine and proline appeared. These aminoacids affect protein surface charge, hydrophilic/hydrophobic properties, conformation density, and structure stability. Conclusions. Changes in high molecular weight glutenin structure, which were caused by the observed mutations, could affect protease ability to effectively hydrolases them, which consequently could affect seed germination rate. Slow seed germination might be one of the mechanisms for survival of wild cereal species Thinopyrum intermedium under unfavorable environmental conditions.Keywords: Thinopyrum intermedium, Glu-1Dх, Glu-1Dy, sequence, point mutations, stop codon.

Intermediate wheatgrass (Thinopyrum intermedium; IWG) is a perennial cereal crop undergoing development for grain production; however, grain yield declines of >75% are often observed after year 2 of the perennial stand and may be linked to soil nutrient depletion. Intercropping IWG with a perennial legume such as alfalfa (Medicago sativa) could benefit nutrient cycling while increasing agroecological diversity. Intermediate wheatgrass was established at five environmentally diverse sites in Minnesota, USA in (1) bi-culture with alfalfa, (2) non-fertilized monoculture and (3) monoculture fertilized annually in the spring with 80 kg N/ha. At northern sites where alfalfa growth was favoured, IWG grain yields were reduced in year 2 by growing IWG in bi-culture with alfalfa, relative to the monoculture systems. Across all sites IWG grain yield decreased by 90% in the non-fertilized monoculture, 80% in the fertilized monoculture and 65% in the bi-culture from year 2 to 4 and plant macronutrient concentrations decreased by 25–70%. In year 4, IWG grain yield was similar or greater in the bi-culture than the fertilized monoculture at three of the five sites and alfalfa biomass was correlated positively with grain yield, harvest index and nutrient uptake in the year 4 bi-culture. Chemical-nitrogen fertilization increased grain yields in year 2 but did not mitigate the decline in yields as stands aged. Intermediate wheatgrass in the bi-culture had similar yields and nutrient uptake and lower yield declines than the chemically fertilized stand at sites where alfalfa growth was maintained throughout the life of the stand.

A comparative study of wheat–wheatgrass substituted cultivars and lines resistant to leaf rust developed by the Agricultural Research Institute for Southeast Regions (Multi 6R, Belyanka, Favorit, Voevoda, Lebedushka) and Samara Agricultural Research Institute (Tulaikovskaya 5, Tulaikovskaya 10, Tulaikovskaya 100, Tulaikovskaya Zolotistaya) breeding was conducted. A complex analysis using molecular cytogenetic (C-differential banding, fluorescent (FISH) and genomic (GISH) in situ hybridization), molecular (PLUG markers), and biochemical (electrophoretic analysis of gliadins) markers demonstrated that they have a substitition of wheat chromosome 6D by the chromosomes 6Agi and 6Agi2 belonging to the J(=E) Agropyron intermedium (Host) Beauv (=Thinopyrum intermedium (Host) Barkworth & D.R. Dewey) subgenome. In spite of the fact that the chromosomes 6Agi and 6Agi2 differ in the C-banding pattern and demonstrated minor differences in the blocks of gliadin components, they had the identical pattern of pSc119.2 and pAs1 probe distribution and conjugated between themselves with insignificant disturbance. Thus, it was demonstrated that 6Agi and 6Agi2 are homologous chromosomes; however, the question about allelism of their leaf rust resistance genes between themselves requires special studies. Nevertheless, using STS and SCAR markers and taking into account the type of reaction to Puccinia triticina, their non-allelism to the Lr9, Lr19, Lr24, Lr29, Lr38, and Lr47 genes was established. It was revealed that the 6Agi and 6Agi2 chromosomes have a different level of transmission in hybrid F2 populations depending on the hybrid combination gene background.

Wheat streak mosaic virus (WSMV) is an important yield constraint of wheat (Triticum aestivum L., AABBDD) throughout the world. Currently, effective WSMV resistance of wheat origin is reported in only one case the Wsm2 gene; excellent disease resistances have been identified in the wild gene pool of Thinopyrum group. Transfer of desirable genes from wild sources has long been a fruitful practice in widening the genetic base of wheat, and Thinopyrum group have enormous genetic variability. WMSV may be controlled by improving agronomic conditions or by applying chemicals (herbicides/miticides), transgenic, or natural resistance sources. Nonetheless, among the available control measures, genetic or naturel resistance to WSMV offers an enduring solution for disease control. This chapter focuses on diverse wheat–Thinopyrum intermedium breeding lines with potential to address WSMV resistance in wheat. The T. intermedium–derived alien chromatin in these hybrid lines carried the Wsm1 gene associated with WSMV resistance in field. The chromosomal translocation was confirmed as 4Ai#2S.4DL with genomic in situ hybridization. Furthermore, for marker-assisted selection breeding, tightly linked PCR-based marker to the alien chromatin was used, and it detected the presence of 4Ai#2S.4DL chromatin. These wheat–T. intermedium recombinant lines increase our arsenal to combat the disease and may allow the development of agronomically desirable WSMV-resistant wheat cultivars.

Intermediate wheatgrass (IWG, Thinopyrum intermedium, (Host) Barkworth &amp, D.R. Dewey subsp. intermedium, 2n = 6x = 42) is a novel perennial crop currently undergoing domestication efforts. It offers remarkable ecosystem services and yields higher relative to other perennial grain crops. While IWG is mostly resistant to Fusarium head blight (FHB), identifying genomic regions associated with resistance will help protect the crop from potential disease epidemics. An IWG biparental population of 108 individuals was developed by crossing parents differing in their response to FHB and bacterial leaf streak (BLS). The population was screened for disease reaction over three years using isolates collected from IWG plants in St. Paul, Minnesota, USA. Linkage maps representing the 21 IWG chromosomes were constructed from 4622 Single Nucleotide Polymorphism (SNP) markers, with one SNP at every 0.74 cM. Interval mapping identified 15 quantitative trait loci (QTL) associated with FHB resistance and 11 with BLS resistance. Models with two or three QTL combinations reduced FHB disease severity by up to 15%, and BLS by up to 17%. When markers associated with FHB resistance were used as cofactors in genomic selection models, trait predictive ability improved by 24&ndash, 125%. These genomic regions and genetic markers associated with FHB and BLS resistance can also be used to safeguard annual cereal grains through gene introgression and selective breeding.

Allohexaploid (2n = 6x = 42) intermediate wheatgrass (Thinopyrum intermedium), abbreviated IWG, is an outcrossing perennial grass belonging to the tertiary gene pool of wheat. Perenniality would be valuable option for grain production, but attempts to introgress this complex trait from wheat-Thinopyrum hybrids have not been commercially successful. Efforts to breed IWG itself as a dual-purpose forage and grain crop have demonstrated useful progress and applications, but grain yields are significantly less than wheat. Therefore, genetic and physical maps have been developed to accelerate domestication of IWG. Herein, these maps were used to identify quantitative trait loci (QTLs) and candidate genes associated with IWG grain production traits in a family of 266 full-sib progenies derived from two heterozygous parents, M26 and M35. Transgressive segregation was observed for 17 traits related to seed size, shattering, threshing, inflorescence capacity, fertility, stem size, and flowering time. A total of 111 QTLs were detected in 36 different regions using 3826 genotype-by-sequence markers in 21 linkage groups. The most prominent QTL had a LOD score of 15 with synergistic effects of 29% and 22% over the family means for seed retention and percentage of naked seeds, respectively. Many QTLs aligned with one or more IWG gene models corresponding to 42 possible domestication orthogenes including the wheat Q and RHT genes. A cluster of seed-size and fertility QTLs showed possible alignment to a putative Z self-incompatibility gene, which could have detrimental grain-yield effects when genetic variability is low. These findings elucidate pathways and possible hurdles in the domestication of IWG.

Perennial crops have been proposed as a more sustainable alternative to annual crops, because they have extended growing seasons, continuous ground cover, reduced nutrient leakage, and sequester more carbon in the soils than annual crops. One example is intermediate wheatgrass (Thinopyrum intermedium), a perennial crop that has been used as a cool-season forage throughout the USA and Canada and also across its native range in Eurasia. Since the 1980's, intermediate wheatgrass has been under domestication to improve seed fertility and grain yield. Commercial products are being sold under the trade name Kernza, owned by The Land Institute, located in Salina, Kansas, USA. This review provides a comprehensive framework about the physical and biological aspects involving the water and carbon cycles in Kernza plants. The main aspects we highlight here are based on previous findings regarding Kernza: i) the ability of maintaining a relatively high water-use efficiency throughout the whole growing season, which is beneficial to mitigate water stress, representing an important physiological mean to acclimate under severe, unfavorable weather conditions, and ii) its higher evapotranspiration (ET) and net carbon uptake rates, particularly when compared to annual counterparts. Only a thorough multifaceted assessment of the repercussion for carbon and water fluxes of a shift from annual crops to Kernza will allow assessing the perspectives of such novel perennial crop to support food security and a number of ecosystem services, particularly under future climates.

A wheat line 14-569 developed from a hybrid of common wheat cv. Chuannong 10 with the wheat–Thinopyrum intermedium partial amphiploid TAI7045 exhibits stable double spikelets per node character in the lower middle portion of a spike, is highly resistant to stripe rust, and produces kernels with a higher 1000-kernel weight than those of its wheat parent. Chromosome counting and an analysis of meiosis demonstrated that line 14-569 is a disomic addition line with 2n = 44, 22II. Sequential genomic in situ hybridization and non-denaturing fluorescence in situ hybridization analyses indicated that one pair of St-genomic chromosomes from Th. intermedium was added to the wheat complement and that the wheat complement included one pair of 1RS/1BL wheat–rye translocation chromosomes. PCR-based landmark unique gene molecular marker analysis revealed that the added pair of chromosomes comprised 3St chromosomes, which suggested that the added 3St chromosomes from Th. intermedium might carry the genes to control the high yield-related characters and stripe rust resistance. This addition line will potentially be useful resource for improvements in wheat yields and stripe rust resistance.

Among the pathogenic complex of the common wheat there is a causal agent of leaf and stem diseases: (rusts and powdery mildew), which has economic importan ce for most of the Russian regions. We propose a methodological approach for confirmation of effec tiveness of wheat genes conferring broad-spectrum resistance to different types of fungal diseases, which integrates traditional field tests and current genotyping techniques with molecular DNA markers. The pro posed approach includes the following steps: 1) evaluating of wheat genotypes using molecular DNA markers; 2) field tests for genotypes; 3) confirma tion of effective ness of broad-spectrum resistance genes by matching field scores and data from mole cular markers. A protocol has been proposed for geno typing varieties and wheat lines using markers linked to Lr16/Sr23, Lr24/ Sr24, Lr19/ Sr25, Lr26/Sr31/Yr9/Pm8, Lr37/Sr38/Yr17 and the gene Lr34(=Sr57/Yr18/Pm38) with a pleiotropic effect. Markers to the genes Lr6Ai#2/Sr6Ai#2/Pm6Ai#2 transferred from Thinopyrum intermedium into the genomes of Russian wheat varieties and markers to a new translocation from Aegi lops spel toides with a group of genes, designated as LrAsp7/SrAsp7/PmAsp7, were tested. When evaluating the contribution of the genes (including genes located on the alien translocations) to the formation of common wheat resistance to fungal diseases, it is necessary to use an extended sample, including genotypes carrying the same group of genes in a different genetic background. In addition, recommendations are given on the terms and frequency of monitoring of fungal diseases in Western Siberia, depending on the pathogen. The methodological approach proposed in the article can be used for identification and evaluation of the efficacy of the genes determining protection of wheat from fungal diseases. This approach is applicable in the investigation of genetic collections consisting of isogenic lines, sources and donors of resistance genes, as well as in the development of wheat genotypes using marker-assisted selection.

Intermediate wheatgrass [Thinopyrum intermedium (Host) Barkworth & D. R. Dewey] plant introductions (PI) have played a critical role in the development of improved intermediate wheatgrass cultivars. The objective of this study was to characterize a large number of intermediate wheatgrass populations over its native range for dry matter yield (DMY), crude protein (CP), in vitro true digestibility (IVTD), neutral detergent fibre (NDF), and rhizome development and genetic diversity and structure to identify superior populations and possible geographical regions in which to collect. Analysis of molecular variance using 643 AFLP bands partitioned 31% of the total genetic variation among these populations with 69% variance detected within populations. Bayesian cluster analysis identified two large groups designated as Asian and European under K = 2. In general, European accessions had greater DMY than did Asian accessions. Observed trends were towards greater CP values in populations with Asian ancestry. Selection index values above zero were observed in populations with European ancestry, and the exception was the late-maturing (208 days to bloom) populations with Asian ancestry. Plant populations 383 551, 383 561, 401 161, 401 173, 440 021, 578 692 (cv. Tegmar) and 598 740 had rhizome scores greater than 7.0. Despite significant differences detected between populations (ΦST = 0.3136; P

New molecular markers were developed for targeting Thinopyrum intermedium 1St#2 chromosome, and novel FISH probe representing the terminal repeats was produced for identification of Thinopyrum chromosomes. Thinopyrum intermedium has been used as a valuable resource for improving the disease resistance and yield potential of wheat. A wheat-Th. intermedium ssp. trichophorum chromosome 1St#2 substitution and translocation has displayed superior grain protein and wet gluten content. With the aim to develop a number of chromosome 1St#2 specific molecular and cytogenetic markers, a high throughput, low-cost specific-locus amplified fragment sequencing (SLAF-seq) technology was used to compare the sequences between a wheat-Thinopyrum 1St#2 (1D) substitution and the related species Pseudoroegneria spicata (St genome, 2n = 14). A total of 5142 polymorphic fragments were analyzed and 359 different SLAF markers for 1St#2 were predicted. Thirty-seven specific molecular markers were validated by PCR from 50 randomly selected SLAFs. Meanwhile, the distribution of transposable elements (TEs) at the family level between wheat and St genomes was compared using the SLAFs. A new oligo-nucleotide probe named Oligo-pSt122 from high SLAF reads was produced for fluorescence in situ hybridization (FISH), and was observed to hybridize to the terminal region of 1St#L and also onto the terminal heterochromatic region of Th. intermedium genomes. The genome-wide markers and repetitive based probe Oligo-pSt122 will be valuable for identifying Thinopyrum chromosome segments in wheat backgrounds.

Exploitation of host resistance is important for controlling powdery mildew of wheat (Triticum aestivum L.). In this study, a wheat-Thinopyrum intermedium introgression line, designated WE99, conferred seedling resistance to 47 of 49 Blumeria graminis f. sp. tritici isolates. Genetic analysis demonstrated that the resistance segregation deviated significantly from a single gene Mendelian ratio. However, marker analysis indicated that only a single recessive resistance gene, temporarily designated PmWE99, conferred powdery mildew resistance (Pm). PmWE99 was mapped to chromosome arm 2BS and linked to the three simple-sequence repeat markers Gwm148, Gwm271, and Barc55. Using race spectrum analysis, PmWE99 was shown to be significantly different from the documented genes Pm42 and MlIW170 located on chromosome arm 2BS and, thus, appeared to be a new Pm gene. Examination of the genotype frequencies in the F2:3 families showed that a genetic variation in the PmWE99 interval that favored the transmission of the WE99 allele could be the cause of the deviated segregation. Further investigation revealed that the abnormal segregation only occurred at the PmWE99 interval and was not common at other loci in this population. Identification of PmWE99 will increase the diversity of the Pm genes for wheat improvement.

Cytogenetic analysis and array-based SNP genotyping of wheat– Th. intermedium introgression lines allowed identification of 634 chromosome-specific SNP markers across all twenty-one chromosomes of Th. intermedium (StJ r J vs , 2 n = 6 x = 42). Thinopyrum intermedium (2n = 6x = 42, StJrJvs) is one of the most promising reservoirs of useful genes including tolerance to abiotic stresses, perenniality and disease resistance not available in the cultivated bread wheat. The transfer of genetic diversity from wild species to wheat offers valuable responses to the effects of climate change. The new array-based single-nucleotide polymorphism (SNP) marker technology provides cheap and easy-to-use molecular markers for marker-assisted selection (MAS) in wheat breeding programmes. Here, we focus on the generation of a new chromosome-specific SNP marker set that can be used to characterize and identify the Th. intermedium chromosomes or chromosome segments transferred into wheat. A progressive investigation of marker development was conducted using 187 various newly developed wheat–Th. intermedium introgression lines and the Axiom® Wheat-Relative Genotyping array. We employed molecular cytogenetic techniques to clarify the genome constitution of the Th. intermedium parental lines and validated 634 chromosome-specific SNPs. Our data confirmed the allohexaploid nature of Th. intermedium and demonstrated that the St genome-specific GISH signal and markers are present at the centromeric regions of chromosomes 1Jvs, 2Jvs, 3Jvs and 7Jvs. The SNP markers presented here will be introduced into current wheat improvement programmes, offering a significant speed-up in wheat breeding and making it possible to deal with the transfer of the full genetic potential of Th. intermedium into wheat.

Synthesized oligonucleotides (oligos) can be used as effective probes similar to plasmid clones for chromosome identification in fluorescence in situ hybridization (FISH) analysis, making oligo FISH a simpler and more efficient molecular cytogenetic technique for studying plants. In this study, multiplex oligonucleotide probes, including pSc119.2-1, pAs1-4, (GAA)10, (AAC)6, and pTa71, were combined and used in FISH to identify chromosomes in common wheat, Thinopyrum intermedium, and a wheat – Th. intermedium amphiploid TE256-1. In comparison with general FISH probes, signals generated by the multiplex probes were more abundant, colorful, and characteristic. Combining the results of genomic in situ hybridization (GISH) with FISH, Th. intermedium chromosomes and alien chromosomes in TE256-1 could be classified and identified more precisely, especially the J- and Js-genome chromosomes. Moreover, based on the FISH results using multiplex probes, more structural variations in wheat chromosomes of TE256-1 were detected. The results indicated that multiplex oligo probes would have a wide range of application prospects in the creation and identification of wheat – Th. intermedium germplasms.

portuguesA ferrugem linear causada por Puccinia striiformis e uma das doencas mais destrutivas do trigo no mundo. A linhagem CH5389 e derivada do cruzamento de trigo com Thinopyrum intermedium e confere resistencia a ferrugem linear. Analises geneticas de individuos da populacao F2 e familia F2:3 obtida a partir do cruzamento entre CH5389 e trigo comum suscetivel revelaram que a resistencia a ferrugem linear na linhagem CH5389 foi controlada por um unico gene dominante, designado YrCH5389. Oito marcadores polimorficos SSR e EST-PCR no cromossomo 3AL foram identificados na populacao F2 de CH5389/Taichung29. O gene YrCH5389 foi delimitado pelos marcadores EST BE405348 e SSR Xwmc388 no cromossomo 3AL com distâncias geneticas de 2,2 e 4,6 cM, respectivamente. Analises genomicas comparativas demonstraram que regioes genomicas ortologas do gene YrCH5389 compreendem 990 kb em arroz, 640 kb em braquipodio e 890 kb em sorgo. Com base nas localizacoes dos marcadores, o gene de resistencia foi localizado no cromossomo 3AL-0.85-1.00. Como nao ha genes oficialmente nomeados de resistencia a ferrugem linear no cromossomo 3AL, o YrCH5389 deve ser designado como um gene novo de resistencia. Esses marcadores de ligacao podem ser uteis para a selecao assistida de genotipos de trigo resistentes a ferrugem linear. Palavras-chave: ferrugem linear; analise genetica; gene de resistencia; analise genomicas comparativas; selecao assistida por marcado EnglishStripe rust, caused by Puccinia striiformis is one of the most destructive diseases of wheat worldwide. CH5389 is a wheat-Thinopyrum intermedium derived line conferring stripe rust resistance. Genetic analyses of seedlings of F2 populations and F2:3 families developed by crossing CH5389 and susceptible common wheat revealed that stripe rust resistance in CH5389 was controlled by a single dominant gene that was designated YrCH5389. Eight SSR and EST-PCR polymorphic markers on chromosome 3AL were identified in F2 population of CH5389/Taichung29. The YrCH5389 was flanked by EST marker BE405348 and SSR marker Xwmc388 on chromosome 3AL with genetic distances of 2.2 and 4.6 cM, respectively. Comparative genomic analysis demonstrated that the orthologous genomic region of YrCH5389 covered 990 kb in rice, 640 kb in Brachypodium, and 890 kb in sorghum. Based on the locations of the markers, the resistance gene was located to chromosome deletion bin 3AL-0.85-1.00. Because there are no officially named stripe rust resistance genes on the 3AL chromosome, the YrCH5389 should be designated as a new resistance gene. These linkage markers could be useful for marker-assisted selection in wheat resistance breeding. Key words: stripe rust; genetic analysis; resistance gene; comparative genomic analysis; marker assisted selection

Key message Genome-wide introgressions of Thinopyrum bessarabicum into wheat resulted in 12 recombinant lines. Cytological and molecular techniques allowed mapping of 1150 SNP markers across all seven chromosomes of the J genome. Abstract Thinopyrum bessarabicum (2n = 2x = 14, JJ) is an important source for new genetic variation for wheat improvement due to its salinity tolerance and disease resistance. Its practical utilisation in wheat improvement can be facilitated through development of genome-wide introgressions leading to a variety of different wheat–Th . bessarabicum translocation lines. In this study, we report the generation of 12 such wheat–Th . bessarabicum recombinant lines, through two different crossing strategies, which were characterized using sequential single colour and multi-colour genomic in situ hybridization (sc-GISH and mc-GISH), multi-colour fluorescent in situ hybridization (mc-FISH) and single nucleotide polymorphic (SNP) DNA markers. We also detected 13 lines containing different Th. bessarabicum chromosome aberrations through sc-GISH. Through a combination of molecular and cytological analysis of all the 25 lines containing Th. bessarabicum recombinants and chromosome aberrations we were able to physically map 1150 SNP markers onto seven Th. bessarabicum J chromosomes which were divided into 36 segmental blocks. Comparative analysis of the physical map of Th. bessarabicum and the wheat genome showed that synteny between the two species is highly conserved at the macro-level and confirmed that Th. bessarabicum contains the 4/5 translocation also present in the A genome of wheat. These wheat–Th . bessarabicum recombinant lines and SNP markers provide a useful genetic resource for wheat improvement with the latter having a wider impact as a tool for detection of introgressions from other Thinopyrum species containing the J or a closely-related genome such as Thinopyrum intermedium (JrJrJvsJvsStSt) and Thinopyrum elongatum (EeEe), respectively. Electronic supplementary material The online version of this article (10.1007/s00122-017-3009-y) contains supplementary material, which is available to authorized users.

The wheat – Thinopyrum intermedium derived line Z4 has displayed novel and effective stripe rust resistance for over 40 years. This study aimed to precisely identify the chromosome constitution of line Z4 and determine the stripe rust resistance contribution using multicolor fluorescent in situ hybridization (FISH) and molecular marker analysis. The results indicated that the Z4 line (2n = 44) contained two pairs of non-Robertsonian translocations without the 3A chromosomes of wheat. FISH karyotypes of F3 progenies derived from crosses between Z4 and MY11 indicated that the transmission of the translocated chromosomes appeared normal and the number of wheat chromosomes 3A and 3D frequently varied. The FISH signal distribution of a new repetitive probe, named Oligo-3A1, confirmed the physical breakage points on chromosome 3AL incorporated into translocated chromosomes. PLUG markers revealed the breakage points on chromosomes 3A, 7JS, and 3D invloved in the translocated chromosomes, and they were designated as T3DS-3AS.3AL-7JSS and T3AL-7JSS.7JSL. Stripe rust resistances surveys indicated that the proximal region of 7JSS or 7JSL may confer the resistance at the adult plant stage. The precise characterization of the chromosome complements of wheat – Th. intermedium Z4 and derived progenies has demonstrated the importance of combining cytogenetic and molecular approaches in the genomics era for further wheat genetic manipulation and breeding purposes.

YrSM139-1B maybe a new gene for effective resistance to stripe rust and useful flanking markers for marker-assisted selection were developed. Stripe rust, caused by Puccinia striiformis f. sp. tritici, is an important foliar disease of wheat. Two dominant stripe rust resistant genes YrSM139-1B and YrSM139-2D were pyramided in bread wheat cultivar Shaanmai 139; one from wild emmer and the other from Thinopyrum intermedium. Three near-isogenic F7:8 line pairs (contrasting RILs), N122-1013R/S, N122-185R/S, and N122-1812R/S, independently derived from different F2 plants and differing at the YrSM139-1B locus were generated from the cross Shaanmai 139 × Hu 901-19 through marker-assisted selection. A large F2:3 population from cross N122-1013R × N122-1013S tested for stripe rust response and subjected to analysis with markers in the 1BS10-0.5 bin region using SSR expressed sequence tags (EST) and site-specific sequence markers developed from the 90 K Illumina iSelect SNP array. Five EST–STS markers and four allele-specific PCR markers were mapped to the YrSM139-1B region. The 30.5 cM genetic map for YrSM139-1B consisted of nine markers, two of which were closer to YrSM139-1B than Xgwm273, which was used in producing the contrasting RIL pairs. Race response data and allelism tests showed that YrSM139-1B is different from Yr10, Yr15, and Yr24/26/CH42.

Thinopyrum intermedium, which has many useful traits, is valuable for wheat breeding. A new wheat-Thinopyrum addition line, SN100109, was developed from the progeny of common wheat cultivar Yannong 15 and Th. intermedium. It was resistant to most races of Blumeria graminis f. sp tritici (Bgt), which caused powdery mildew in wheat, and its reactions were different from the reactions of gene Pm40 and Pm43. Genomic in situ hybridization (GISH) and molecular marker analysis were used to identify the genomic composition of SN100109. GISH results showed that SN100109 was a wheat-Th. intermedium disomic addition line containing one pair of J chromosomes, and the resistance gene was located on the alien additional chromosomes of SN100109. And four molecular markers BE425942, BF482714, Xgdm93 and BV679214 which were assigned to homologous group 2, were specific molecular markers of the additional chromosomes. All the results indicated that SN100109 contained one pair of 2J chromosomes. SN100109 can be used as a novel germplasm source for introducing powdery mildew resistance genes to wheat in breeding programs.

Intermediate wheatgrass (Thinopyrum intermedium (Host) Barkworth & D.R. Dewey), a segmental autoallohexaploid (2n = 6x = 42), is not only an important forage crop but also a valuable gene reservoir for wheat (Triticum aestivum L.) improvement. Throughout the scientific literature, there continues to be disagreement as to the origin of the different genomes in intermediate wheatgrass. Genotypic data obtained from newly developed EST-SSR primers derived from the putative progenitor diploid species Pseudoroegneria spicata (Pursh) Á. Löve (St genome), Thinopyrum bessarabicum (Savul. & Rayss) Á. Löve (J = Jb= Eb), and Thinopyrum elongatum (Host) D. Dewey (E = Je= Ee) indicate that the V genome of Dasypyrum (Coss. & Durieu) T. Durand is not one of the three genomes in intermediate wheatgrass. Based on all available information in the literature and findings in this study, the genomic designation of intermediate wheatgrass should be changed to JvsJrSt, where Jvsand Jrrepresent ancestral genomes of present-day Jbof Th. bessarabicum and Jeof Th. elongatum, with Jvsbeing more ancient. Furthermore, the information suggests that the St genome in intermediate wheatgrass is most similar to the present-day St found in diploid species of Pseudoroegneria from Eurasia.

Wheatgrass Thinopyrum intermedium (Host), or Agropyron intermedium (Host) (2n = 42; genome formula, EEEstEstStSt) is one of the most valuable sources of highly effective resistance genes in wheat breeding. Bread wheat cultivars Tulaikovskaya 5, Tulaikovskaya 10, and Tulaikovskaya 100 derived from wheat—Thinopyrum crosses are highly resistant to leaf rust and powdery mildew, and moderately resistant to stem and yellow rust. C-banding, in situ hybridization, and assays with PLUG and SSR markers have demonstrated that wheat chromosome 6D in all three cultivars is substituted by the Th. intermedium homoeologous chromosome, 6Ai. This chromosome was designated 6Ai#2, because it differs from the earlier described homoeologous chromosome 6Ai#1. In situ hybridization with Pseudoroegneria spicata and Dasypyrum villosum genomic DNAs has allowed chromosome 6Ai#2 to be assigned to the E (=J) subgenome. Chromosome 6Ai#2 remained intact over long-term breeding efforts. Tests of leaf rust resistance of in F2 and F3 populations from a cross of a leaf rust–susceptible cultivar with Tulaikovskaya 10 has demonstrated that chromosome 6Ai#2 carries at least one gene locus for leaf rust resistance. This locus was designated Lr6Ai#2. It must be different from the known gene locus Lr38, located on the long arm of chromosome 7Ai#2 and now present in many translocation variants in bread wheat. The effect of a chromosome from the Th. intermedium subgenome E on resistance to powdery mildew, stem and yellow rust in Tulaikovskaya 5, Tulaikovskaya 10, and Tulaikovskaya 100 is discussed.