Mapping and characterization of rust resistance genes Lr53 and Yr35 introgressed from Aegilops species.

KEY MESSAGE: The rust resistance genes Lr53 and Yr35 were introgressed into bread wheat from Aegilops longissima or Aegilops sharonensis or their S-genome containing species and mapped to the telomeric region of chromosome arm 6BS. Wheat leaf and stripe rusts are damaging fungal diseases of wheat worldwide. Breeding for resistance is a sustainable approach to control these two foliar diseases. In this study, we used SNP analysis, sequence comparisons, and cytogenetic assays to determine that the chromosomal segment carrying Lr53 and Yr35 was originated from Ae.longissima or Ae. sharonensis or their derived species. In seedling tests, Lr53 conferred strong resistance against all five Chinese Pt races tested, and Yr35 showed effectiveness against Pst race CYR34 but susceptibility to race CYR32. Using a large population (3892 recombinant gametes) derived from plants homozygous for the ph1b mutation obtained from the cross 98M71 × CSph1b, both Lr53 and Yr35 were successfully mapped to a 6.03-Mb telomeric region of chromosome arm 6BS in the Chinese Spring reference genome v1.1. Co-segregation between Lr53 and Yr35 was observed within this large mapping population. Within the candidate region, several nucleotide-binding leucine-rich repeat genes and protein kinases were identified as candidate genes. Marker pku6B3127 was completely linked to both genes and accurately predicted the absence or presence of alien segment harboring Lr53 and Yr35 in 87 tetraploid and 149 hexaploid wheat genotypes tested. We developed a line with a smaller alien segment (< 6.03 Mb) to reduce any potential linkage drag and demonstrated that it conferred resistance levels similar to those of the original donor parent 98M71. The newly developed introgression line and closely linked PCR markers will accelerate the deployment of Lr53 and Yr35 in wheat breeding programs.

Dissecting the molecular basis of spike traits by integrating gene regulatory networks and genetic variation in wheat.

Spike architecture influences both grain weight and grain number per spike, which are the two major components of grain yield in bread wheat (Triticum aestivum L.). However, the complex wheat genome and the influence of various environmental factors pose challenges in mapping the causal genes that affect spike traits. Here, we systematically identified genes involved in spike trait formation by integrating information on genomic variation and gene regulatory networks controlling young spike development in wheat. We identified 170 loci that are responsible for variations in spike length, spikelet number per spike, and grain number per spike through genome-wide association study and meta-QTL analyses. We constructed gene regulatory networks for young inflorescences at the double ridge stage and the floret primordium stage, in which the spikelet meristem and the floret meristem are predominant, respectively, by integrating transcriptome, histone modification, chromatin accessibility, eQTL, and protein-protein interactome data. From these networks, we identified 169 hub genes located in 76 of the 170 QTL regions whose polymorphisms are significantly associated with variation in spike traits. The functions of TaZF-B1, VRT-B2, and TaSPL15-A/D in establishment of wheat spike architecture were verified. This study provides valuable molecular resources for understanding spike traits and demonstrates that combining genetic analysis and developmental regulatory networks is a robust approach for dissection of complex traits.

Chemical-tag-based semi-annotated metabolomics facilitates gene identification and specialized metabolic pathway elucidation in wheat.

The importance of metabolite modification and species-specific metabolic pathways has long been recognized. However, linking the chemical structure of metabolites to gene function in order to explore the genetic and biochemical basis of metabolism has not yet been reported in wheat (Triticum aestivum). Here, we profiled metabolic fragment enrichment in wheat leaves and consequently applied chemical-tag-based semi-annotated metabolomics in a genome-wide association study in accessions of wheat. The studies revealed that all 1,483 quantified metabolites have at least one known functional group whose modification is tailored in an enzyme-catalyzed manner and eventually allows efficient candidate gene mining. A Triticeae crop-specific flavonoid pathway and its underlying metabolic gene cluster were elucidated in further functional studies. Additionally, upon overexpressing the major effect gene of the cluster TraesCS2B01G460000 (TaOMT24), the pathway was reconstructed in rice (Oryza sativa), which lacks this pathway. The reported workflow represents an efficient and unbiased approach for gene mining using forward genetics in hexaploid wheat. The resultant candidate gene list contains vast molecular resources for decoding the genetic architecture of complex traits and identifying valuable breeding targets and will ultimately aid in achieving wheat crop improvement.

Chemoselective Coatings of GL13K Antimicrobial Peptides for Dental Implants.

Dental implant-associated infection is a clinical challenge which poses a significant healthcare and socio-economic burden. To overcome this issue, developing antimicrobial surfaces, including antimicrobial peptide coatings, has gained great attention. Different physical and chemical routes have been used to obtain these biofunctional coatings, which in turn might have a direct influence on their bioactivity and functionality. In this study, we present a silane-based, fast, and efficient chemoselective conjugation of antimicrobial peptides (Cys-GL13K) to coat titanium implant surfaces. Comprehensive surface analysis was performed to confirm the surface functionalization of as-prepared and mechanically challenged coatings. The antibacterial potency of the evaluated surfaces was confirmed against both Streptococcus gordonii and Streptococcus mutans, the primary colonizers and pathogens of dental surfaces, as demonstrated by reduced bacteria viability. Additionally, human dental pulp stem cells demonstrated long-term viability when cultured on Cys-GL13K-grafted titanium surfaces. Cell functionality and antimicrobial capability against multi-species need to be studied further; however, our results confirmed that the proposed chemistry for chemoselective peptide anchoring is a valid alternative to traditional site-unspecific anchoring methods and offers opportunities to modify varying biomaterial surfaces to form potent bioactive coatings with multiple functionalities to prevent infection.

Cloning of the wheat leaf rust resistance gene Lr47 introgressed from Aegilops speltoides.

Leaf rust, caused by Puccinia triticina Eriksson (Pt), is one of the most severe foliar diseases of wheat. Breeding for leaf rust resistance is a practical and sustainable method to control this devastating disease. Here, we report the identification of Lr47, a broadly effective leaf rust resistance gene introgressed into wheat from Aegilops speltoides. Lr47 encodes a coiled-coil nucleotide-binding leucine-rich repeat protein that is both necessary and sufficient to confer Pt resistance, as demonstrated by loss-of-function mutations and transgenic complementation. Lr47 introgression lines with no or reduced linkage drag are generated using the Pairing homoeologous1 mutation, and a diagnostic molecular marker for Lr47 is developed. The coiled-coil domain of the Lr47 protein is unable to induce cell death, nor does it have self-protein interaction. The cloning of Lr47 expands the number of leaf rust resistance genes that can be incorporated into multigene transgenic cassettes to control this devastating disease.

Identification and validation of new quantitative trait loci for spike-related traits in two RIL populations.

Wheat (Triticum aestivum L.) is one of the most important cereal crops for ensuring food security worldwide. Identification of major quantitative trait loci (QTL) for spike-related traits is important for improvement of yield potential in wheat breeding. In this study, by using the wheat 55K single nucleotide polymorphism (SNP) array and diversity array technology (DArT), two recombinant inbred line populations derived from crosses avocet/chilero and avocet/huites were used to map QTL for kernel number per spike (KNS), total spikelet number per spike (TSS), fertile spikelet number per spike (FSS), and spike compactness (SC). Forty-two QTLs were identified on chromosomes 2A (4), 2B (3), 3A (2), 3B (7), 5A (11), 6A (4), 6B, and 7A (10), explaining 3.13-21.80% of the phenotypic variances. Twelve QTLs were detected in multi-environments on chromosomes 2A, 3B (2), 5A (4), 6A (3), 6B, and 7A, while four QTL clusters were detected on chromosomes 3A, 3B, 5A, and 7A. Two stable and new QTL clusters, QKns/Tss/Fss/SC.haust-5A and QKns/Tss/Fss.haust-7A, were detected in the physical intervals of 547.49-590.46 Mb and 511.54-516.15 Mb, accounting for 7.53-14.78% and 7.01-20.66% of the phenotypic variances, respectively. High-confidence annotated genes for QKns/Tss/Fss/SC.haust-5A and QKns/Tss/Fss.haust-7A were more highly expressed in spike development. The results provide new QTL and molecular markers for marker-assisted breeding in wheat. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01401-4.

Quality traits analysis of 153 wheat lines derived from CIMMYT and China.

In order to understand the difference of quality for Chinese and CIMMYT wheat varieties (lines), we selected 153 wheat germplasm from both China and CIMMYT to explore the contribution relationship of different allelic variation combinations to wheat quality through genotyping and phenotyping, including grain hardness, polyphenol oxidase (PPO) activity, lipoxygenase (LOX) activity, yellow pigment (YP) content and protein content. In terms of flour milling quality, Chinese wheat varieties were mainly carrying Pina-D1a/Pinb-D1b, accounting for 32.0% of the total tested varieties, while the CIMMYT wheat lines were mainly carrying Pina-D1b/Pinb-D1a with 45.8% of the total collection. The distribution frequencies of subunit 1/2* and 5 + 10 were 47.0% and 42.5%, respectively, in CIMMYT varieties, however they were only 31.4% and 13.7% respectively of the Chinese wheat tested varieties. In addition, the proportion of phytoene synthase (PSY) allele, PPO allele and LOX active allele were roughly the same between Chinese and CIMMYT varieties. Based on the present study, we found that Pina gene had a greater impact on grain hardness value than Pinb gene; The influence of PPO-A1 gene on polyphenol oxidase activity was more significant than PPO-D1 gene. The high protein content of varieties mostly containing hardness genes and 1/2*/5 + 10 subunit combinations. Based on the present study, we found that the quality gene distribution of Chinese and CIMMYT varieties was quite different, for instance, the high-quality HMW-GS subunits of Chinese varieties were lower than CIMMYT lines. It will be much useful for Chinese wheat breeders to develop good quality wheat variety by crossing with 3 good strong gluten CIMMYT wheat lines by molecular marker-assisted selection.

Centromere Plasticity With Evolutionary Conservation and Divergence Uncovered by Wheat 10+ Genomes.

Centromeres (CEN) are the chromosomal regions that play a crucial role in maintaining genomic stability. The underlying highly repetitive DNA sequences can evolve quickly in most eukaryotes, and promote karyotype evolution. Despite their variability, it is not fully understood how these widely variable sequences ensure the homeostasis of centromere function. In this study, we investigated the genetics and epigenetics of CEN in a population of wheat lines from global breeding programs. We captured a high degree of sequences, positioning, and epigenetic variations in the large and complex wheat CEN. We found that most CENH3-associated repeats are Cereba element of retrotransposons and exhibit phylogenetic homogenization across different wheat lines, but the less-associated repeat sequences diverge on their own way in each wheat line, implying specific mechanisms for selecting certain repeat types as functional core CEN. Furthermore, we observed that CENH3 nucleosome structures display looser wrapping of DNA termini on complex centromeric repeats, including the repositioned CEN. We also found that strict CENH3 nucleosome positioning and intrinsic DNA features play a role in determining centromere identity among different lines. Specific non-B form DNAs were substantially associated with CENH3 nucleosomes for the repositioned centromeres. These findings suggest that multiple mechanisms were involved in the adaptation of CENH3 nucleosomes that can stabilize CEN. Ultimately, we proposed a remarkable epigenetic plasticity of centromere chromatin within the diverse genomic context, and the high robustness is crucial for maintaining centromere function and genome stability in wheat 10+ lines as a result of past breeding selections.

Genetic analysis of stripe rust resistance in the common wheat line Kfa/2*Kachu under a Chinese rust environment.

KEY MESSAGE: We mapped a new race-specific seedling stripe rust resistance gene on wheat chromosome 5BL and a new APR locus QYr.hazu-2BS from CIMMYT wheat line Kfa/2*Kachu. Breeding resistant wheat (Triticum aestivum) varieties is the most economical and efficient way to manage wheat stripe rust, but requires the prior identification of new resistance genes and development of associated molecular markers for marker-assisted selection. To map stripe rust resistance loci in wheat, we used a recombinant inbred line population generated by crossing the stripe rust-resistant parent 'Kfa/2*Kachu' and the susceptible parent 'Apav#1'. We employed genotyping-by-sequencing and bulked segregant RNA sequencing to map a new race-specific seedling stripe rust resistance gene, which we designated YrK, to wheat chromosome arm 5BL. TraesCS5B02G330700 encodes a receptor-like kinase and is a high-confidence candidate gene for YrK based on virus-induced gene silencing results and the significant induction of its expression 24 h after inoculation with wheat stripe rust. To assist breeding, we developed functional molecular markers based on the polymorphic single nucleotide polymorphisms in the coding sequence region of YrK. We also mapped four adult plant resistance (APR) loci to wheat chromosome arms 1BL, 2AS, 2BS and 4AL. Among these APR loci, we determined that QYr.hazu-1BL and QYr.hazu-2AS are allelic to the known pleiotropic resistance gene Lr46/Yr29/Pm39 and the race-specific gene Yr17, respectively. However, QYr.hazu-2BS is likely a new APR locus, for which we converted closely linked SNP polymorphisms into breeder-friendly Kompetitive allele-specific PCR (KASP) markers. In the present study, we provided new stripe rust resistance locus/gene and molecular markers for wheat breeder to develop rust-resistant wheat variety.

Slow stripe rusting in Chinese wheat Jimai 44 conferred by Yr29 in combination with a major QTL on chromosome arm 6AL.

KEY MESSAGE: YrJ44, a more effective slow rusting gene than Yr29, was localized to a 3.5-cM interval between AQP markers AX-109373479 and AX-109563479 on chromosome 6AL. "Slow rusting" (SR) is a type of adult plant resistance (APR) that can provide non-specific durable resistance to stripe rust in wheat. Chinese elite wheat cultivar Jimai 44 (JM44) has maintained SR to stripe rust in China since its release despite exposure to a changing and variable pathogen population. An F2:6 population comprising 295 recombinant inbred lines (RILs) derived from a cross between JM44 and susceptible cultivar Jimai 229 (JM229) was used in genetic analysis of the SR. The RILs and parental lines were evaluated for stripe rust response in five field environments and genotyped using the Affymetrix Wheat55K SNP array and 13 allele-specific quantitative PCR-based (AQP) markers. Two stable QTL on chromosome arms 1BL and 6AL were identified by inclusive composite interval mapping. The 1BL QTL was probably the pleiotropic gene Lr46/Yr29/Sr58. QYr.nwafu-6AL (hereafter named YrJ44), mapped in a 3.5-cM interval between AQP markers AX-109373479 and AX-109563479, was more effective than Yr29 in reducing disease severity and relative area under the disease progress curve (rAUDPC). RILs harboring both YrJ44 and Yr29 displayed levels of SR equal to the resistant parent JM44. The AQP markers linked with YrJ44 were polymorphic and significantly correlated with stripe rust resistance in a panel of 1,019 wheat cultivars and breeding lines. These results suggested that adequate SR resistance can be obtained by combining YrJ44 and Yr29 and the AQP markers can be used in breeding for durable stripe rust resistance.

Deciphering genetic basis of developmental and agronomic traits by integrating high-throughput optical phenotyping and genome-wide association studies in wheat.

Dissecting the genetic basis of complex traits such as dynamic growth and yield potential is a major challenge in crops. Monitoring the growth throughout growing season in a large wheat population to uncover the temporal genetic controls for plant growth and yield-related traits has so far not been explored. In this study, a diverse wheat panel composed of 288 lines was monitored by a non-invasive and high-throughput phenotyping platform to collect growth traits from seedling to grain filling stage and their relationship with yield-related traits was further explored. Whole genome re-sequencing of the panel provided 12.64 million markers for a high-resolution genome-wide association analysis using 190 image-based traits and 17 agronomic traits. A total of 8327 marker-trait associations were detected and clustered into 1605 quantitative trait loci (QTLs) including a number of known genes or QTLs. We identified 277 pleiotropic QTLs controlling multiple traits at different growth stages which revealed temporal dynamics of QTLs action on plant development and yield production in wheat. A candidate gene related to plant growth that was detected by image traits was further validated. Particularly, our study demonstrated that the yield-related traits are largely predictable using models developed based on i-traits and provide possibility for high-throughput early selection, thus to accelerate breeding process. Our study explored the genetic architecture of growth and yield-related traits by combining high-throughput phenotyping and genotyping, which further unravelled the complex and stage-specific contributions of genetic loci to optimize growth and yield in wheat.

Asymmetric gene expression and cell-type-specific regulatory networks in the root of bread wheat revealed by single-cell multiomics analysis.

BACKGROUND: Homoeologs are defined as homologous genes resulting from allopolyploidy. Bread wheat, Triticum aestivum, is an allohexaploid species with many homoeologs. Homoeolog expression bias, referring to the relative contribution of homoeologs to the transcriptome, is critical for determining the traits that influence wheat growth and development. Asymmetric transcription of homoeologs has been so far investigated in a tissue or organ-specific manner, which could be misleading due to a mixture of cell types. RESULTS: Here, we perform single nuclei RNA sequencing and ATAC sequencing of wheat root to study the asymmetric gene transcription, reconstruct cell differentiation trajectories and cell-type-specific gene regulatory networks. We identify 22 cell types. We then reconstruct cell differentiation trajectories that suggest different origins between epidermis/cortex and endodermis, distinguishing bread wheat from Arabidopsis. We show that the ratio of asymmetrically transcribed triads varies greatly when analyzing at the single-cell level. Hub transcription factors determining cell type identity are also identified. In particular, we demonstrate that TaSPL14 participates in vasculature development by regulating the expression of BAM1. Combining single-cell transcription and chromatin accessibility data, we construct the pseudo-time regulatory network driving root hair differentiation. We find MYB3R4, REF6, HDG1, and GATAs as key regulators in this process. CONCLUSIONS: Our findings reveal the transcriptional landscape of root organization and asymmetric gene transcription at single-cell resolution in polyploid wheat.

QTL cluster analysis and marker development for kernel traits based on DArT markers in spring bread wheat (Triticum aestivum L.).

Genetic dissection of yield component traits including kernel characteristics is essential for the continuous improvement in wheat yield. In the present study, one recombinant inbred line (RIL) F6 population derived from a cross between Avocet and Chilero was used to evaluate the phenotypes of kernel traits of thousand-kernel weight (TKW), kernel length (KL), and kernel width (KW) in four environments at three experimental stations during the 2018-2020 wheat growing seasons. The high-density genetic linkage map was constructed with the diversity arrays technology (DArT) markers and the inclusive composite interval mapping (ICIM) method to identify the quantitative trait loci (QTLs) for TKW, KL, and KW. A total of 48 QTLs for three traits were identified in the RIL population on the 21 chromosomes besides 2A, 4D, and 5B, accounting for 3.00%-33.85% of the phenotypic variances. Based on the physical positions of each QTL, nine stable QTL clusters were identified in the RILs, and among these QTL clusters, TaTKW-1A was tightly linked to the DArT marker interval 3950546-1213099, explaining 10.31%-33.85% of the phenotypic variances. A total of 347 high-confidence genes were identified in a 34.74-Mb physical interval. TraesCS1A02G045300 and TraesCS1A02G058400 were among the putative candidate genes associated with kernel traits, and they were expressed during grain development. Moreover, we also developed high-throughput kompetitive allele-specific PCR (KASP) markers of TaTKW-1A, validated in a natural population of 114 wheat varieties. The study provides a basis for cloning the functional genes underlying the QTL for kernel traits and a practical and accurate marker for molecular breeding.

The pathway of melatonin biosynthesis in common wheat (Triticum aestivum).

Melatonin (Mel) is a multifunctional biomolecule found in both animals and plants. In plants, the biosynthesis of Mel from tryptophan (Trp) has been delineated to comprise of four consecutive reactions. However, while the genes encoding these enzymes in rice are well characterized no systematic evaluation of the overall pathway has, as yet, been published for wheat. In the current study, the relative contents of six Mel-pathway-intermediates including Trp, tryptamine (Trm), serotonin (Ser), 5-methoxy tryptamine (5M-Trm), N-acetyl serotonin (NAS) and Mel, were determined in 24 independent tissues spanning the lifetime of wheat. These studies indicated that Trp was the most abundant among the six metabolites, followed by Trm and Ser. Next, the candidate genes expressing key enzymes involved in the Mel pathway were explored by means of metabolite-based genome-wide association study (mGWAS), wherein two TDC genes, a T5H gene and one SNAT gene were identified as being important for the accumulation of Mel pathway metabolites. Moreover, a 463-bp insertion within the T5H gene was discovered that may be responsible for variation in Ser content. Finally, a ASMT gene was found via sequence alignment against its rice homolog. Validations of these candidate genes were performed by in vitro enzymatic reactions using proteins purified following recombinant expression in Escherichia coli, transient gene expression in tobacco, and transgenic approaches in wheat. Our results thus provide the first comprehensive investigation into the Mel pathway metabolites, and a swift candidate gene identification via forward-genetics strategies, in common wheat.

Stripe rust and leaf rust resistance in CIMMYT wheat line "Mucuy" is conferred by combinations of race-specific and adult-plant resistance loci.

Developing wheat varieties with durable resistance is a core objective of the International Maize and Wheat Improvement Center (CIMMYT) and many other breeding programs worldwide. The CIMMYT advanced wheat line "Mucuy" displayed high levels of resistance to stripe rust (YR) and leaf rust (LR) in field evaluations in Mexico and several other countries. To determine the genetic basis of YR and LR resistance, 138 F5 recombinant inbred lines (RILs) derived from the cross of Apav#1× Mucuy were phenotyped for YR responses from 2015 to 2020 at field sites in India, Kenya, and Mexico, and LR in Mexico. Seedling phenotyping for YR and LR responses was conducted in the greenhouse in Mexico using the same predominant races as in field trials. Using 12,681 polymorphic molecular markers from the DArT, SNP, and SSR genotyping platforms, we constructed genetic linkage maps and QTL analyses that detected seven YR and four LR resistance loci. Among these, a co-located YR/LR resistance loci was identified as Yr29/Lr46, and a seedling stripe rust resistance gene YrMu was mapped on the 2AS/2NS translocation. This fragment also conferred moderate adult plant resistance (APR) under all Mexican field environments and in one season in Kenya. Field trial phenotyping with Lr37-virulent Puccinia triticina races indicated the presence of an APR QTL accounting for 18.3-25.5% of the LR severity variation, in addition to a novel YR resistance QTL, QYr.cim-3DS, derived from Mucuy. We developed breeder-friendly KASP and indel molecular markers respectively for Yr29/Lr46 and YrMu. The current study validated the presence of known genes and identified new resistance loci, a QTL combination effect, and flanking markers to facilitate accelerated breeding for genetically complex, durable rust resistance.

Genetic Analysis of Adult Plant Resistance to Stripe Rust in Common Wheat Cultivar "Pascal".

Wheat stripe rust is an important foliar disease that affects the wheat yield globally. Breeding for resistant wheat varieties is one of the most economically and environmentally effective ways to control this disease. The common wheat (Triticum aestivum L.) cultivar "Pascal" exhibited susceptibility to stripe rust at the seedling stage but it showed high resistance to stripe rust at the adult plant stage over 20 years in Gansu, a hotspot of the disease in northwestern China. To understand the genetic mechanism of stripe rust resistance in this cultivar, a 55K SNP array was used to analyze the two parents and the 220 recombinant inbred lines (RILs) derived from the cross of "Huixianhong" × "Pascal." We detected three new stripe rust adult plant resistance (APR) quantitative trait locus (QTL) contributed by Pascal, viz. QYr.gaas-1AL, QYr.gaas-3DL, and QYr.gaas-5AS, using the inclusive composite interval mapping method. They were flanked by SNP markers AX-111218361-AX-110577861, AX-111460455-AX-108798599, and AX-111523523-AX-110028503, respectively, and explained the phenotypic variation ranging from 11.0 to 23.1%. Bulked segregant exome capture sequencing (BSE-Seq) was used for fine mapping of QYr.gaas-1AL and selection of candidate genes, TraesCS1A02G313700, TraesCS1A02G313800, and TraesCS1A02G314900 for QYr.gaas-1AL. KASP markers BSE-1A-12 and HXPA-3D for QYr.gaas-1AL and QYr.gaas-3DL were developed for breeders to develop durable stripe rust-resistant wheat varieties.

Identification and Characterization of Resistance Loci to Wheat Leaf Rust and Stripe Rust in Afghan Landrace "KU3067".

Leaf rust and stripe rust are important wheat diseases worldwide causing significant losses where susceptible varieties are grown. Resistant cultivars offer long-term control and reduce the use of hazardous chemicals, which can be detrimental to both human health and the environment. Land races have been a valuable resource for mining new genes for various abiotic and biotic stresses including wheat rusts. Afghan wheat landrace "KU3067" displayed high seedling infection type (IT) for leaf rust and low IT for stripe rust; however, it displayed high levels of field resistance for both rusts when tested for multiple seasons against the Mexican rust isolates. This study focused on identifying loci-conferring seedling resistance to stripe rust, and also loci-conferring adult plant resistance (APR) against the Mexican races of leaf rust and stripe rust. A backcrossed inbred line (BIL) population advanced to the BC1F5 generation derived from the cross of KU3067 and Apav (triple rust susceptible line) was used for both, inheritance and QTL mapping studies. The population and parents were genotyped with Diversity Arrays Technology-genotyping-by-sequencing (DArT-Seq) and phenotyped for leaf rust and stripe rust response at both seedling and adult plant stages during multiple seasons in Mexico with relevant pathotypes. Mapping results identified an all-stage resistance gene for stripe rust, temporarily designated as YrKU, on chromosome 7BL. In total, six QTL-conferring APR to leaf rust on 1AS, 2AL, 4DL, 6BL, 7AL, and 7BL, and four QTL for stripe rust resistance on 1BS, 2AL, 4DL, and 7BL were detected in the analyses. Among these, pleiotropic gene Lr67/Yr46 on 4DL with a significantly large effect is the first report in an Afghan landrace-conferring resistance to both leaf and stripe rusts. QLr.cim-7BL/YrKU showed pleiotropic resistance to both rusts and explained 7.5-17.2 and 12.6-19.3% of the phenotypic variance for leaf and stripe rusts, respectively. QYr.cim-1BS and QYr.cim-2AL detected in all stripe environments with phenotypic variance explained (PVE) 12.9-20.5 and 5.4-12.5%, and QLr.cim-6BL are likely to be new. These QTL and their closely linked markers will be useful for fine mapping and marker-assisted selection (MAS) in breeding for durable resistance to multiple rust diseases.

Production of Conjoined Transgenic and Edited Barley and Wheat Plants for Nud Genes Using the CRISPR/SpCas9 System.

The Nudum (Nud) gene controls the caryopsis type of cereal crops by regulating lipid biosynthetic pathways. Based on the HvNud sequence and its homologous gene sequences in wheat, a conserved sgRNA was designed to obtain the mutants from the barley variety "Vlamingh" and the wheat variety "Fielder" via Agrobacterium-mediated transformation. A total of 19 and 118 transgenic plants were obtained, and 11 and 61 mutant plants were identified in T0 transgenic plants in barley and wheat after PCR-RE detection, and the editing efficiencies of the targeted gene were 57.9 and 51.7% in barley and wheat, respectively. The grain shape of the barley mutants was naked. Five different combinations of mutations for wheat TaNud genes were identified in the T0 generation, and their homozygous-edited plants were obtained in the T1 generation. Interestingly, the conjoined plants in which one plant has different genotypes were first identified. The different tillers in an individual T0 plant showed independent transgenic or mutant events in both barley and wheat, and the different genotypes can stably inherit into T1 generation, indicating that the T0 transgenic plants were the conjoined type. In addition, we did not find any off-target mutations in both barley and wheat. A candidate method for detecting putative-edited wheat plants was suggested to avoid losing mutations in this investigation. This study provides not only materials for studying the function of the Nud gene in barley and wheat but also a system for detecting the mutants in wheat.

Pleiotropic effect analysis and marker development for grain zinc and iron concentrations in spring wheat.

Wheat (Triticum aestivum L.) is one of the main food crops in the world and a primary source of zinc (Zn) and iron (Fe) in the human body. The genetic mechanisms underlying related traits have been clarified, thereby providing a molecular theoretical foundation for the development of germplasm resources. In this study, a total of 23,536 high-quality DArT markers was used to map quantitative trait loci (QTL) of grain Zn (GZn) and grain Fe (GFe) concentrations in recombinant inbred lines crossed by Avocet/Chilero. A total of 17 QTLs was located on chromosomes 1BL, 2BL, 3BL, 4AL, 4BS, 5AL, 5DL, 6AS, 6BS, 6DS, and 7AS accounting for 0.38-16.62% of the phenotypic variance. QGZn.haust-4AL, QGZn.haust-7AS.1, and QGFe.haust-6BS were detected on chromosomes 4AL, 6BS, and 7AS, accounting for 10.63-16.62% of the phenotypic variance. Four stable QTLs, QGZn.haust-4AL, QGFe.haust-1BL, QGFe.haust-4AL, and QGFe.haust-5DL, were located on chromosomes 1BL, 4AL, and 5DL. Three pleiotropic effects loci for GZn and GFe concentrations were located on chromosomes 1BL, 4AL, and 5DL. Two high-throughput Kompetitive Allele Specific PCR markers were developed by closely linking single-nucleotide polymorphisms on chromosomes 4AL and 5DL, which were validated by a germplasm panel. Therefore, it is the most important that quantitative trait loci and KASP marker for grain zinc and iron concentrations were developed for utilizing in marker-assisted breeding and biofortification of wheat grain in breeding programs.