Pm57 from Aegilops searsii encodes a tandem kinase protein and confers wheat powdery mildew resistance.

Powdery mildew is a devastating disease that affects wheat yield and quality. Wheat wild relatives represent valuable sources of disease resistance genes. Cloning and characterization of these genes will facilitate their incorporation into wheat breeding programs. Here, we report the cloning of Pm57, a wheat powdery mildew resistance gene from Aegilops searsii. It encodes a tandem kinase protein with putative kinase-pseudokinase domains followed by a von Willebrand factor A domain (WTK-vWA), being ortholog of Lr9 that mediates wheat leaf rust resistance. The resistance function of Pm57 is validated via independent mutants, gene silencing, and transgenic assays. Stable Pm57 transgenic wheat lines and introgression lines exhibit high levels of all-stage resistance to diverse isolates of the Bgt fungus, and no negative impacts on agronomic parameters are observed in our experimental set-up. Our findings highlight the emerging role of kinase fusion proteins in plant disease resistance and provide a valuable gene for wheat breeding.

Identification of candidate genes for adult plant stripe rust resistance transferred from Aegilops ventricosa 2NvS into wheat via fine mapping and transcriptome analysis.

KEY MESSAGE: An adult plant gene for resistance to stripe rust was narrowed down to the proximal one-third of the 2NvS segment translocated from Aegilops ventricosa to wheat chromosome arm 2AS, and based on the gene expression analysis, two candidate genes were identified showing a stronger response at the adult plant stage compared to the seedling stage. The 2NvS translocation from Aegilops ventricosa, known for its resistance to various diseases, has been pivotal in global wheat breeding for more than three decades. Here, we identified an adult plant resistance (APR) gene in the 2NvS segment in wheat line K13-868. Through fine mapping in a segregating near-isogenic line (NIL) derived population of 6389 plants, the candidate region for the APR gene was narrowed down to between 19.36 Mb and 33 Mb in the Jagger reference genome. Transcriptome analysis in NILs strongly suggested that this APR gene conferred resistance to stripe rust by triggering plant innate immune responses. Based on the gene expression analysis, two disease resistance-associated genes within the candidate region, TraesJAG2A03G00588940 and TraesJAG2A03G00590140, exhibited a stronger response to Puccinia striiformis f. sp. tritici (Pst) infection at the adult plant stage than at the seedling stage, indicating that they could be potential candidates for the resistance gene. Additionally, we developed a co-dominant InDel marker, InDel_31.05, for detecting this APR gene. Applying this marker showed that over one-half of the wheat varieties approved in 2021 and 2022 in Sichuan province, China, carry this gene. Agronomic trait evaluation of NILs indicated that the 2NvS segment effectively mitigated the negative effects of stripe rust on yield without affecting other important agronomic traits. This study provided valuable insights for cloning and breeding through the utilization of the APR gene present in the 2NvS segment.

Characterization and validation of TaAGL66, a gene related to fertility conversion of wheat in the presence of Aegilops kotschyi cytoplasm.

MAIN CONCLUSION: TaAGL66, a MADS-box transcription factor highly expressed in fertile anthers of KTM3315A, regulates anther and/or pollen development, as well as male fertility in wheat with Aegilops kotschyi cytoplasm. Male sterility, as a string of sophisticated biological processes in higher plants, is commonly regulated by transcription factors (TFs). Among them, MADS-box TFs are mainly participated in the processes of floral organ formation and pollen development, which are tightly related to male sterility, but they have been little studied in the reproductive development in wheat. In our study, TaAGL66, a gene that was specifically expressed in spikes and highly expressed in fertile anthers, was identified by RNA sequencing and the expression profiles data of these genes, and qRT-PCR analyses, which was localized to the nucleus. Silencing of TaAGL66 under fertility condition in KTM3315A, a thermo-sensitive male sterile line with Ae. kotschyi cytoplasm, displayed severe fertility reduction, abnormal anther dehiscence, defective pollen development, decreased viability, and low seed-setting. It can be concluded that TaAGL66 plays an important role in wheat pollen development in the presence of Ae. kotschyi cytoplasm, providing new insights into the utilization of male sterility.

Chromosome stability of synthetic Triticum turgidum-Aegilops umbellulata hybrids.

BACKGROUND: Unreduced gamete formation during meiosis plays a critical role in natural polyploidization. However, the unreduced gamete formation mechanisms in Triticum turgidum-Aegilops umbellulata triploid F1 hybrid crosses and the chromsome numbers and compostions in T. turgidum-Ae. umbellulata F2 still not known. RESULTS: In this study, 11 T.turgidum-Ae. umbellulata triploid F1 hybrid crosses were produced by distant hybridization. All of the triploid F1 hybrids had 21 chromosomes and two basic pathways of meiotic restitution, namely first-division restitution (FDR) and single-division meiosis (SDM). Only FDR was found in six of the 11 crosses, while both FDR and SDM occurred in the remaining five crosses. The chromosome numbers in the 127 selfed F2 seeds from the triploid F1 hybrid plants of 10 crosses (no F2 seeds for STU 16) varied from 35 to 43, and the proportions of euploid and aneuploid F2 plants were 49.61% and 50.39%, respectively. In the aneuploid F2 plants, the frequency of chromosome loss/gain varied among genomes. The chromosome loss of the U genome was the highest (26.77%) among the three genomes, followed by that of the B (22.83%) and A (11.81%) genomes, and the chromosome gain for the A, B, and U genomes was 3.94%, 3.94%, and 1.57%, respectively. Of the 21 chromosomes, 7U (16.54%), 5 A (3.94%), and 1B (9.45%) had the highest loss frequency among the U, A, and B genomes. In addition to chromosome loss, seven chromosomes, namely 1 A, 3 A, 5 A, 6 A, 1B, 1U, and 6U, were gained in the aneuploids. CONCLUSION: In the aneuploid F2 plants, the frequency of chromosome loss/gain varied among genomes, chromsomes, and crosses. In addition to variations in chromosome numbers, three types of chromosome translocations including 3UL·2AS, 6UL·1AL, and 4US·6AL were identified in the F2 plants. Furthermore, polymorphic fluorescence in situ hybridization karyotypes for all the U chromosomes were also identified in the F2 plants when compared with the Ae. umbellulata parents. These results provide useful information for our understanding the naturally occurred T. turgidum-Ae. umbellulata amphidiploids.

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.

Discrepancy of flowering time between genetically close sublineages of Aegilops umbellulata Zhuk.

Aegilops umbellulata Zhuk., a wild diploid wheat-related species, has been used as a genetic resource for several important agronomic traits. However, its genetic variations have not been comprehensively studied. We sequenced RNA from 114 accessions of Ae. umbellulata to evaluate DNA polymorphisms and phenotypic variations. Bayesian clustering and phylogenetic analysis based on SNPs detected by RNA sequencing revealed two divergent lineages, UmbL1 and UmbL2. The main differences between them were in the sizes of spikes and spikelets, and culm diameter. UmbL1 is divided into two sublineages, UmbL1e and UmbL1w. These genetic differences corresponded to geographic distributions. UmbL1e, UmbL1w, and UmbL2 are found in Turkey, Iran/Iraq, and Greece, respectively. Although UmbL1e and UmbL1w were genetically similar, flowering time and other morphological traits were more distinct between these sublineages than those between the lineages. This discrepancy can be explained by the latitudinal and longitudinal differences in habitats. Specifically, latitudinal clines of flowering time were clearly observed in Ae. umbellulata, strongly correlated with solar radiation in the winter season. This observation implies that latitudinal differences are a factor in differences in the flowering times of Ae. umbellulata. Differences in flowering time could influence other morphological differences and promote genetic divergence between sublineages.

Wheat powdery mildew resistance gene Pm13 encodes a mixed lineage kinase domain-like protein.

Wheat powdery mildew is one of the most destructive diseases threatening global wheat production. The wild relatives of wheat constitute rich sources of diversity for powdery mildew resistance. Here, we report the map-based cloning of the powdery mildew resistance gene Pm13 from the wild wheat species Aegilops longissima. Pm13 encodes a mixed lineage kinase domain-like (MLKL) protein that contains an N-terminal-domain of MLKL (MLKL_NTD) domain in its N-terminus and a C-terminal serine/threonine kinase (STK) domain. The resistance function of Pm13 is validated by mutagenesis, gene silencing, transgenic assay, and allelic association analyses. The development of introgression lines with significantly reduced chromosome segments of Ae. longissima encompassing Pm13 enables widespread deployment of this gene into wheat cultivars. The cloning of Pm13 may provide valuable insights into the molecular mechanisms underlying Pm13-mediated powdery mildew resistance and highlight the important roles of kinase fusion proteins (KFPs) in wheat immunity.

Variation of TaMyb10 and their function on grain color and pre-harvest sprouting resistance of wheat.

Pre-harvest sprouting (PHS) is a significant threat to global food security due to its association with losses in both yield and quality. Among the genes involved in PHS resistance in wheat, PHS-3D (TaMyb10-D) plays a crucial role. Here, we characterized the sequence variations of TaMyb10 genes in 416 bread wheat and 302 Aegilops tauschii accessions. Within TaMyb10-A sequences, we identified a deletion ranging from 214 to 305 bp in the signal and amino acid coding region, present in 61.3% of the accessions. Similarly, 79.3% of the TaMyb10-B sequences within the third exon region exhibited a 19 bp deletion. Additionally, 40.8% of the accessions lacked the 2.4 Mb fragment (in/del mutations) on Chr3D, where TaMyb10-D/PHS-3D was located. Interestingly, the geographical distribution of accessions showed little correlation with the divergence of TaMyb10. TaMyb10-A-IIIDele, TaMyb10-B-IVDele, and TaMyb10-D-VDele genotypes were prevalent in wheat populations across continents. Despite their structural variations, the five distinct protein types exhibited comparable ability to bind the promoters of downstream genes in the flavonoid and ABA pathways, such as CHS, DFR, and NCED. Furthermore, the combination of TaMyb10 homologs was significantly associated with grain color and germination percentages. Accessions exclusively harboring TaMyb10-D displayed red seed color and reduced germination percentages, indicating the predominant role of TaMyb10-D compared to TaMyb10-A and TaMyb10-B. This comprehensive investigation enhances our understanding of the structural variations and functional divergence of TaMyb10, providing valuable insights and resources for improving PHS resistance in wheat.

The R2R3 MYB gene TaMYB305 positively regulates anther and pollen development in thermo-sensitive male-sterility wheat with Aegilops kotschyi cytoplasm.

MAIN CONCLUSION: The loss of TaMYB305 function down-regulated the expression of jasmonic acid synthesis pathway genes, which may disturb the jasmonic acid synthesis, resulting in abnormal pollen development and reduced fertility. The MYB family, as one of the largest transcription factor families found in plants, regulates plant development, especially the development of anthers. Therefore, it is important to identify potential MYB transcription factors associated with pollen development and to study its role in pollen development. Here, the transcripts of an R2R3 MYB gene TaMYB305 from KTM3315A, a thermo-sensitive cytoplasmic male-sterility line with Aegilops kotschyi cytoplasm (K-TCMS) wheat, was isolated. Quantitative real-time PCR (qRT-PCR) and promoter activity analysis revealed that TaMYB305 was primarily expressed in anthers. The TaMYB305 protein was localized in the nucleus, as determined by subcellular localization analysis. Our data demonstrated that silencing of TaMYB305 was related to abnormal development of stamen, including anther indehiscence and pollen abortion in KAM3315A plants. In addition, TaMYB305-silenced plants exhibited alterations in the transcriptional levels of genes involved in the synthesis of jasmonic acid (JA), indicating that TaMYB305 may regulate the expression of genes related to JA synthesis and play an important role during anther and pollen development of KTM3315A. These results provide novel insight into the function and molecular mechanism of R2R3-MYB genes in pollen development.

Transcriptome analysis in Aegilops tauschii unravels further insights into genetic control of stripe rust resistance.

MAIN CONCLUSION: The Aegilops tauschii resistant accession prevented the pathogen colonization by controlling the sugar flow and triggering the hypersensitive reaction. This study suggested that NBS-LRRs probably induce resistance through bHLH by controlling JA- and SA-dependent pathways. Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst) is one of wheat's most destructive fungal diseases that causes a severe yield reduction worldwide. The most effective and economically-friendly strategy to manage this disease is genetic resistance which can be achieved through deploying new and effective resistance genes. Aegilops tauschii, due to its small genome and co-evolution with Pst, can provide detailed information about underlying resistance mechanisms. Hence, we used RNA-sequencing approach to identify the transcriptome variations of two contrasting resistant and susceptible Ae. tauschii accessions in interaction with Pst and differentially expressed genes (DEGs) for resistance to stripe rust. Gene ontology, pathway analysis, and search for functional domains, transcription regulators, resistance genes, and protein-protein interactions were used to interpret the results. The genes encoding NBS-LRR, CC-NBS-kinase, and phenylalanine ammonia-lyase, basic helix-loop-helix (bHLH)-, basic-leucine zipper (bZIP)-, APETALA2 (AP2)-, auxin response factor (ARF)-, GATA-, and LSD-like transcription factors were up-regulated exclusively in the resistant accession. The key genes involved in response to salicylic acid, amino sugar and nucleotide sugar metabolism, and hypersensitive response contributed to plant defense against stripe rust. The activation of jasmonic acid biosynthesis and starch and sucrose metabolism pathways under Pst infection in the susceptible accession explained the colonization of the host. Overall, this study can fill the gaps in the literature on host-pathogen interaction and enrich the Ae. tauschii transcriptome sequence information. It also suggests candidate genes that could guide future breeding programs attempting to develop rust-resistant cultivars.

Exploring Aegilops caudata: A Comprehensive Study of the CslF6 Gene and ß-Glucan.

In the quest for sustainable and nutritious food sources, exploration of ancient grains and wild relatives of cultivated cereals has gained attention. Aegilops caudata, a wild wheatgrass species, stands out as a promising genetic resource due to its potential for crop enhancement and intriguing nutritional properties. This manuscript investigates the CslF6 gene sequence and protein structure of Aegilops caudata, employing comparative analysis with other grass species to identify potential differences impacting ß-glucan content. The study involves comprehensive isolation and characterization of the CslF6 gene in Ae. caudata, utilizing genomic sequence analysis, protein structure prediction, and comparative genomics. Comparisons with sequences from diverse monocots reveal evolutionary relationships, highlighting high identities with wheat genomes. Specific amino acid motifs in the CslF6 enzyme sequence, particularly those proximal to key catalytic motifs, exhibit variations among monocot species. These differences likely contribute to alterations in ß-glucan composition, notably impacting the DP3:DP4 ratio, which is crucial for understanding and modulating the final ß-glucan content. The study positions Ae. caudata uniquely within the evolutionary landscape of CslF6 among monocots, suggesting potential genetic divergence or unique functional adaptations within this species. Overall, this investigation enriches our understanding of ß-glucan biosynthesis, shedding light on the role of specific amino acid residues in modulating enzymatic activity and polysaccharide composition.

A platform for whole-genome speed introgression from Aegilops tauschii to wheat for breeding future crops.

Breeding new and sustainable crop cultivars of high yields and desirable traits has been a major challenge for ensuring food security for the growing global human population. For polyploid crops such as wheat, introducing genetic variation from wild relatives of its subgenomes is a key strategy to improve the quality of their breeding pools. Over the past decades, considerable progress has been made in speed breeding, genome sequencing, high-throughput phenotyping and genomics-assisted breeding, which now allows us to realize whole-genome introgression from wild relatives to modern crops. Here, we present a standardized protocol to rapidly introgress the entire genome of Aegilops tauschii, the progenitor of the D subgenome of bread wheat, into elite wheat backgrounds. This protocol integrates multiple modern high-throughput technologies and includes three major phases: development of synthetic octaploid wheat, generation of hexaploid A. tauschii-wheat introgression lines (A-WIs) and homozygosis of the generated A-WIs. Our approach readily generates stable introgression lines in 2 y, thus greatly accelerating the generation of A-WIs and the introduction of desirable genes from A. tauschii to wheat cultivars. These A-WIs are valuable for wheat-breeding programs and functional gene discovery. The current protocol can be easily modified and used for introgressing the genomes of wild relatives to other polyploid crops.

Evolutionary trajectory of organelle-derived nuclear DNAs in the Triticum/Aegilops complex species.

Organelle-derived nuclear DNAs, nuclear plastid DNAs (NUPTs), and nuclear mitochondrial DNAs (NUMTs) have been identified in plants. Most, if not all, genes residing in NUPTs/NUMTs (NUPGs/NUMGs) are known to be inactivated and pseudogenized. However, the role of epigenetic control in silencing NUPGs/NUMGs and the dynamic evolution of NUPTs/NUMTs with respect to organismal phylogeny remain barely explored. Based on the available nuclear and organellar genomic resources of wheat (genus Triticum) and goat grass (genus Aegilops) within Triticum/Aegilops complex species, we investigated the evolutionary fates of NUPTs/NUMTs in terms of their epigenetic silencing and their dynamic occurrence rates in the nuclear diploid genomes and allopolyploid subgenomes. NUPTs and NUMTs possessed similar genomic atlas, including (i) predominantly located in intergenic regions and preferential integration to gene regulation regions and (ii) generating sequence variations in the nuclear genome. Unlike nuclear indigenous genes, the alien NUPGs/NUMGs were associated with repressive epigenetic signals, namely high levels of DNA methylation and low levels of active histone modifications. Phylogenomic analyses suggested that the species-specific and gradual accumulation of NUPTs/NUMTs accompanied the speciation processes. Moreover, based on further pan-genomic analyses, we found significant subgenomic asymmetry in the NUPT/NUMT occurrence, which accumulated during allopolyploid wheat evolution. Our findings provide insight into the dynamic evolutionary fates of organelle-derived nuclear DNA in plants.

Effects of Aegilops longissima chromosome 1Sl on wheat bread-making quality in two types of translocation lines.

KEY MESSAGE: Two wheat-Ae. longissima translocation chromosomes (1BS·1SlL and 1SlS·1BL) were transferred into three commercial wheat varieties, and the new advanced lines showed improved bread-making quality compared to their recurrent parents. Aegilops longissima chromosome 1Sl encodes specific types of gluten subunits that may positively affect wheat bread-making quality. The most effective method of introducing 1Sl chromosomal fragments containing the target genes into wheat is chromosome translocation. Here, a wheat-Ae. longissima 1BS·1SlL translocation line was developed using molecular marker-assisted chromosome engineering. Two types of translocation chromosomes developed in a previous study, 1BS·1SlL and 1SlS·1BL, were introduced into three commercial wheat varieties (Ningchun4, Ningchun50, and Westonia) via backcrossing with marker-assisted selection. Advanced translocation lines were confirmed through chromosome in situ hybridization and genotyping by target sequencing using the wheat 40 K system. Bread-making quality was found to be improved in the two types of advanced translocation lines compared to the corresponding recurrent parents. Furthermore, 1SlS·1BL translocation lines displayed better bread-making quality than 1BS·1SlL translocation lines in each genetic background. Further analysis revealed that high molecular weight glutenin subunit (HMW-GS) contents and expression levels of genes encoding low molecular weight glutenin subunits (LMW-GSs) were increased in 1SlS·1BL translocation lines. Gliadin and gluten-related transcription factors were also upregulated in the grains of the two types of advanced translocation lines compared to the recurrent parents. This study clarifies the impacts of specific glutenin subunits on bread-making quality and provides novel germplasm resources for further improvement of wheat quality through molecular breeding.

Chromosome genomics facilitates the marker development and selection of wheat-Aegilops biuncialis addition, substitution and translocation lines.

The annual goatgrass, Aegilops biuncialis is a rich source of genes with considerable agronomic value. This genetic potential can be exploited for wheat improvement through interspecific hybridization to increase stress resistance, grain quality and adaptability. However, the low throughput of cytogenetic selection hampers the development of alien introgressions. Using the sequence of flow-sorted chromosomes of diploid progenitors, the present study enabled the development of chromosome-specific markers. In total, 482 PCR markers were validated on wheat (Mv9kr1) and Ae. biuncialis (MvGB642) crossing partners, and 126 on wheat-Aegilops additions. Thirty-two markers specific for U- or M-chromosomes were used in combination with GISH and FISH for the screening of 44 Mv9kr1 × Ae. biuncialis BC3F3 genotypes. The predominance of chromosomes 4M and 5M, as well as the presence of chromosomal aberrations, may indicate that these chromosomes have a gametocidal effect. A new wheat-Ae. biuncialis disomic 4U addition, 4M(4D) and 5M(5D) substitutions, as well as several introgression lines were selected. Spike morphology and fertility indicated that the Aegilops 4M or 5M compensated well for the loss of 4D and 5D, respectively. The new cytogenetic stocks represent valuable genetic resources for the introgression of key genes alleles into wheat.

Chromosome-scale assembly of the wild wheat relative Aegilops umbellulata.

Wild wheat relatives have been explored in plant breeding to increase the genetic diversity of bread wheat, one of the most important food crops. Aegilops umbellulata is a diploid U genome-containing grass species that serves as a genetic reservoir for wheat improvement. In this study, we report the construction of a chromosome-scale reference assembly of Ae. umbellulata accession TA1851 based on corrected PacBio HiFi reads and chromosome conformation capture. The total assembly size was 4.25 Gb with a contig N50 of 17.7 Mb. In total, 36,268 gene models were predicted. We benchmarked the performance of hifiasm and LJA, two of the most widely used assemblers using standard and corrected HiFi reads, revealing a positive effect of corrected input reads. Comparative genome analysis confirmed substantial chromosome rearrangements in Ae. umbellulata compared to bread wheat. In summary, the Ae. umbellulata assembly provides a resource for comparative genomics in Triticeae and for the discovery of agriculturally important genes.

Fine mapping of two recessive powdery mildew resistance genes from Aegilops tauschii accession CIae8.

KEY MESSAGE: Two recessive powdery mildew resistance loci pmAeCIae8_2DS and pmAeCIae8_7DS from Aegilops tauschii were mapped and two synthesized hexaploid wheat lines were developed by distant hybridization. Wheat powdery mildew (Pm), one of the worldwide destructive fungal diseases, causes significant yield loss up to 30%. The identification of new Pm resistance genes will enrich the genetic diversity of wheat breeding for Pm resistance. Aegilops tauschii is the ancestor donor of sub-genome D of hexaploid wheat. It provides beneficial genes that can be easily transferred into wheat by producing synthetic hexaploid wheat followed by genetic recombination. We assessed the Pm resistance level of 35 Ae. tauschii accessions from different origins. Accession CIae8 exhibited high Pm resistance. Inheritance analysis and gene mapping were performed using F2 and F2:3 populations derived from the cross between CIae8 and a Pm susceptible accession PI574467. The Pm resistance of CIae8 was controlled by two independent recessive genes. Bulked segregate analysis using a 55 K SNP array revealed the SNPs were mainly enriched into genome regions, i.e. 2DS (13.5-20 Mb) and 7DS (4.0-15.5 Mb). The Pm resistance loci were named as pmAeCIae8_2DS and pmAeCIae8_7DS, respectively. By recombinant screening, we narrowed the pmAeCIae8_2DS into a 370-kb interval flanked by markers CINAU-AE7800 (14.89 Mb) and CINAU-AE20 (15.26 Mb), and narrowed the pmAeCIae8_7DS into a 260-kb interval flanked by markers CINAU-AE58 (4.72 Mb) and CINAU-AE25 (4.98 Mb). The molecular markers closely linked with the resistance loci were developed, and two synthesized hexaploid wheat (SHW) lines were produced. These laid the foundation for cloning of the two resistance loci and for transferring the resistance into common wheat.

Genome-wide identification, phylogenetic and expression pattern analysis of HSF family genes in the Rye (Secale cereale L.).

BACKGROUND: Heat shock factor (HSF), a typical class of transcription factors in plants, has played an essential role in plant growth and developmental stages, signal transduction, and response to biotic and abiotic stresses. The HSF genes families has been identified and characterized in many species through leveraging whole genome sequencing (WGS). However, the identification and systematic analysis of HSF family genes in Rye is limited. RESULTS: In this study, 31 HSF genes were identified in Rye, which were unevenly distributed on seven chromosomes. Based on the homology of A. thaliana, we analyzed the number of conserved domains and gene structures of ScHSF genes that were classified into seven subfamilies. To better understand the developmental mechanisms of ScHSF family during evolution, we selected one monocotyledon (Arabidopsis thaliana) and five (Triticum aestivum L., Hordeum vulgare L., Oryza sativa L., Zea mays L., and Aegilops tauschii Coss.) specific representative dicotyledons associated with Rye for comparative homology mapping. The results showed that fragment replication events modulated the expansion of the ScHSF genes family. In addition, interactions between ScHSF proteins and promoters containing hormone- and stress-responsive cis-acting elements suggest that the regulation of ScHSF expression was complex. A total of 15 representative genes were targeted from seven subfamilies to characterize their gene expression responses in different tissues, fruit developmental stages, three hormones, and six different abiotic stresses. CONCLUSIONS: This study demonstrated that ScHSF genes, especially ScHSF1 and ScHSF3, played a key role in Rye development and its response to various hormones and abiotic stresses. These results provided new insights into the evolution of HSF genes in Rye, which could help the success of molecular breeding in Rye.

Genetic analysis of iron, zinc and grain yield in wheat-Aegilops derivatives using multi-locus GWAS.

BACKGROUND: Wheat is a major staple crop and helps to reduce worldwide micronutrient deficiency. Investigating the genetics that control the concentrations of iron (Fe) and zinc (Zn) in wheat is crucial. Hence, we undertook a comprehensive study aimed at elucidating the genomic regions linked to the contents of Fe and Zn in the grain. METHODS AND RESULTS: We performed the multi-locus genome-wide association (ML-GWAS) using a panel of 161 wheat-Aegilops substitution and addition lines to dissect the genomic regions controlling grain iron (GFeC), and grain zinc (GZnC) contents. The wheat panel was genotyped using 10,825 high-quality SNPs and phenotyped in three different environments (E1-E3) during 2017-2019. A total of 111 marker-trait associations (MTAs) (at p-value < 0.001) were detected that belong to all three sub-genomes of wheat. The highest number of MTAs were identified for GFeC (58), followed by GZnC (44) and yield (9). Further, six stable MTAs were identified for these three traits and also two pleiotropic MTAs were identified for GFeC and GZnC. A total of 1291 putative candidate genes (CGs) were also identified for all three traits. These CGs encode a diverse set of proteins, including heavy metal-associated (HMA), bZIP family protein, AP2/ERF, and protein previously associated with GFeC, GZnC, and grain yield. CONCLUSIONS: The significant MTAs and CGs pinpointed in this current study are poised to play a pivotal role in enhancing both the nutritional quality and yield of wheat, utilizing marker-assisted selection (MAS) techniques.

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.