Two functional CC-NBS-LRR proteins from rye chromosome 6RS confer differential age-related powdery mildew resistance to wheat.

Rye (Secale cereale), a valuable relative of wheat, contains abundant powdery mildew resistance (Pm) genes. Using physical mapping, transcriptome sequencing, barley stripe mosaic virus-induced gene silencing, ethyl methane sulfonate mutagenesis, and stable transformation, we isolated and validated two coiled-coil, nucleotide-binding site and leucine-rich repeat (CC-NBS-LRR) alleles, PmTR1 and PmTR3, located on rye chromosome 6RS from different triticale lines. PmTR1 confers age-related resistance starting from the three-leaf stage, whereas its allele, PmTR3, confers typical all-stage resistance, which may be associated with their differential gene expression patterns. Overexpression in Nicotiana benthamiana showed that the CC, CC-NBS, and CC-LRR fragments of PMTR1 induce cell death, whereas in PMTR3 the CC and full-length fragments perform this function. Luciferase complementation imaging and pull-down assays revealed distinct interaction activities between the CC and NBS fragments. Our study elucidates two novel rye-derived Pm genes and their derivative germplasm resources and provides novel insights into the mechanism of age-related resistance, which can aid the improvement of resistance against wheat powdery mildew.

Unraveling the genetic basis of grain number-related traits in a wheat-Agropyron cristatum introgressed line through high-resolution linkage mapping.

BACKGROUND: Grain number per spike (GNS) is a pivotal determinant of grain yield in wheat. Pubing 3228 (PB3228), a wheat-Agropyron cristatum germplasm, exhibits a notably higher GNS. RESULTS: In this study, we developed a recombinant inbred line (RIL) population derived from PB3228/Gao8901 (PG-RIL) and constructed a high-density genetic map comprising 101,136 loci, spanning 4357.3 cM using the Wheat 660 K SNP array. The genetic map demonstrated high collinearity with the wheat assembly IWGSC RefSeq v1.0. Traits related to grain number and spikelet number per spike were evaluated in seven environments for quantitative trait locus (QTL) analysis. Five environmentally stable QTLs were detected in at least three environments. Among these, two major QTLs, QGns-4A.2 and QGns-1A.1, associated with GNS, exhibited positive alleles contributed by PB3228. Further, the conditional QTL analysis revealed a predominant contribution of PB3228 to the GNS QTLs, with both grain number per spikelet (GNSL) and spikelet number per spike (SNS) contributing to the overall GNS trait. Four kompetitive allele-specific PCR (KASP) markers that linked to QGns-4A.2 and QGns-1A.1 were developed and found to be effective in verifying the QTL effect within a diversity panel. Compared to previous studies, QGns-4A.2 exhibited stability across different trials, while QGns-1A.1 represents a novel QTL. The results from unconditional and conditional QTL analyses are valuable for dissecting the genetic contribution of the component traits to GNS at the individual QTL level and for understanding the genetic basis of the superior grain number character in PB3228. The KASP markers can be utilized in marker-assisted selection for enhancing GNS. CONCLUSIONS: Five environmentally stable QTLs related to grain number and spikelet number per spike were identified. PB3228 contributed to the majority of the QTLs associated with GNS.

Development of novel wheat-rye 6RS small fragment translocation lines with powdery mildew resistance and physical mapping of the resistance gene PmW6RS.

KEY MESSAGE: Novel wheat-rye 6RS small fragment translocation lines with powdery mildew resistance were developed, and the resistance gene PmW6RS was physically mapped onto 6RS-0.58-0.66-bin corresponding to 18.38 Mb in Weining rye. Rye (Secale cereale L., RR) contains valuable genes for wheat improvement. However, most of the rye resistance genes have not been successfully used in wheat cultivars. Identification of new rye resistance genes and transfer of these genes to wheat by developing small fragment translocation lines will make these genes more usable for wheat breeding. In this study, a broad-spectrum powdery mildew resistance gene PmW6RS was localized on rye chromosome arm 6RS using a new set of wheat-rye disomic and telosomic addition lines. To further study and use PmW6RS, 164 wheat-rye 6RS translocation lines were developed by 60Coγ-ray irradiation. Seedling and adult stage powdery mildew resistance analysis showed that 106 of the translocation lines were resistant. A physical map of 6RS was constructed using the 6RS translocation and deletion lines, and PmW6RS was localized in the 6RS-0.58-0.66-bin, flanked by markers X6RS-3 and X6RS-10 corresponding to the physical interval of 50.23-68.61 Mb in Weining rye genome. A total of 23 resistance-related genes were annotated. Nine markers co-segregate with the 6RS-0.58-0.66-bin, which can be used to rapidly trace the 6RS fragment carrying PmW6RS. Small fragment translocation lines with powdery mildew resistance were backcrossed with wheat cultivars, and 39 agronomically acceptable homozygous 6RS small fragment translocation lines were obtained. In conclusion, this study not only provides novel gene source and germplasms for wheat resistance breeding, but also laid a solid foundation for cloning of PmW6RS.

The osmotic stress-activated receptor-like kinase DPY1 mediates SnRK2 kinase activation and drought tolerance in Setaria.

Plant genomes encode many receptor-like kinases (RLKs) that localize to the cell surface and perceive a wide variety of environmental cues to initiate downstream signaling cascades. Whether these RLKs participate in dehydration stress signaling in plants is largely unknown. DROOPY LEAF1 (DPY1), a leucine-rich repeat (LRR)-RLK, was recently shown to regulate plant architecture by orchestrating early brassinosteroid signaling in foxtail millet (Setaria italica). Here, we show that DPY1 is essential for the acclimation of foxtail millet to drought stress. DPY1 can be phosphorylated and activated in response to osmotic stress and is required for more than half of osmotic stress-induced global phosphorylation events, including the phosphorylation of sucrose nonfermenting kinase 2s (SnRK2s), the central kinases involved in osmotic stress. DPY1 acts upstream of STRESS-ACTIVATED PROTEIN KINASE 6 (SAPK6, a subclass I SnRK2) and is required for full SAPK6 activation, thereby allowing regulation of downstream genes to mount a response against drought stress. These signaling events are largely independent of DPY1-mediated brassinosteroid signaling. The DPY1-SAPK6 module is specific to seed plants and is absent in ancestral nonseed plants. Our findings reveal a dehydration stress-activated RLK that plays an indispensable role in osmotic stress signaling and mediates SnRK2 activation at the cell surface.

Identification of major QTLs for yield-related traits with improved genetic map in wheat.

Introduction: Identification of stable major quantitative trait loci (QTLs) for yield-related traits is important for yield potential improvement in wheat breeding. Methods: In the present study, we genotyped a recombinant inbred line (RIL) population using the Wheat 660K SNP array and constructed a high-density genetic map. The genetic map showed high collinearity with the wheat genome assembly. Fourteen yield-related traits were evaluated in six environments for QTL analysis. Results and Discussion: A total of 12 environmentally stable QTLs were identified in at least three environments, explaining up to 34.7% of the phenotypic variation. Of these, QTkw-1B.2 for thousand kernel weight (TKW), QPh-2D.1 (QSl-2D.2/QScn-2D.1) for plant height (PH), spike length (SL) and spikelet compactness (SCN), QPh-4B.1 for PH, and QTss-7A.3 for total spikelet number per spike (TSS) were detected in at least five environments. A set of Kompetitive Allele Specific PCR (KASP) markers were converted based on the above QTLs and used to genotype a diversity panel comprising of 190 wheat accessions across four growing seasons. QPh-2D.1 (QSl-2D.2/QScn-2D.1), QPh-4B.1 and QTss-7A.3 were successfully validated. Compared with previous studies, QTkw-1B.2 and QPh-4B.1 should be novel QTLs. These results provided a solid foundation for further positional cloning and marker-assisted selection of the targeted QTLs in wheat breeding programs.

Development and Identification of a Wheat-Rye Breeding Line for Harmonious Improvement Between Powdery Mildew Resistance and High Yield Potential.

Powdery mildew, caused by Blumeria graminis f. sp. tritici, is a devastating disease that seriously threatens wheat yield and quality. To control this disease, host resistance is the preferred measure. However, wheat breeding is a complex process with elusive exchange and recombination of the traits from their parents. Increased resistance often leads to a decline in other key traits, such as yield and quality. Developing breakthrough germplasms with harmonious powdery mildew resistance and other key breeding traits is attractive in wheat breeding. In this study, we developed an ideal wheat breeding line AL46 that pyramided its hexaploid triticale parent-derived desirable yield traits and its wheat parent-derived powdery mildew resistance gene Pm2. Sequential genomic in situ hybridization (GISH), multicolor GISH, multicolor fluorescence in situ hybridization, and molecular marker analyses revealed that AL46 was a wheat-rye T1RS·1BL translocation line. Genetic analysis combined with function marker detection and sequence alignment were used to confirm that AL46 carried the Pm2 gene. Then, we evaluated the powdery mildew resistance and comprehensive traits of AL46, and just as we designed, AL46 showed harmonious powdery mildew resistance with some key breeding traits. This study not only developed an ideal wheat germplasm resource but also provided a successful example for pyramiding breeding, which could be a promising direction for wheat improvement in the future.

Identification of a Wheat Powdery Mildew Dominant Resistance Gene in the Pm5 Locus for High-Throughput Marker-Assisted Selection.

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), poses a severe threat to wheat yield and quality worldwide. Rapid identification and the accurate transference of effective resistance genes are important to the development of resistant cultivars and the sustainable control of this disease. In the present study, the wheat line AL11 exhibited high levels of resistance to powdery mildew at both the seedling and adult plant stages. Genetic analysis of the AL11 × 'Shixin 733' mapping population revealed that its resistance was controlled by a single dominant gene, tentatively designated PmAL11. Using bulked segregant RNA-Seq and molecular marker analysis, PmAL11 was mapped to the Pm5 locus on chromosome 7B where it cosegregated with the functional marker Pm5e-KASP. Sequence alignment analysis revealed that the Pm5e-homologous sequence in AL11 was identical to the reported recessive gene Pm5e in wheat landrace 'Fuzhuang 30'. It appears that PmAL11 was most probably Pm5e, but it was mediated by a dominant inheritance pattern, so it should provide a valuable resistance resource for both genetic study and wheat breeding. To efficiently use and trace PmAL11 in breeding, a new kompetitive allele-specific PCR marker AL11-K2488 that cosegregated with this gene was developed and confirmed to be applicable in the different wheat backgrounds, thus promoting its use in the marker-assisted selection of PmAL11.

Identification and validation of plant height, spike length and spike compactness loci in common wheat (Triticum aestivum L.).

BACKGROUND: Plant height (PH), spike length (SL) and spike compactness (SCN) are important agronomic traits in wheat due to their strong correlations with lodging and yield. Thus, dissection of their genetic basis is essential for the improvement of plant architecture and yield potential in wheat breeding. The objective of this study was to map quantitative trait loci (QTL) for PH, SL and SCN in a recombinant inbred line (RIL) population derived from the cross 'PuBing3228 × Gao8901' (PG-RIL) and to evaluate the potential values of these QTL to improve yield. RESULTS: In the current study, Five, six and ten stable QTL for PH, SL, and SCN, respectively, were identified in at least two individual environments. Five major QTL QPh.cas-5A.3, QPh.cas-6A, QSl.cas-6B.2, QScn.cas-2B.2 and QScn.cas-6B explained 5.58-25.68% of the phenotypic variation. Notably, two, three and three novel stable QTL for PH, SL and SCN were identified in this study, which could provide further insights into the genetic factors that shape PH and spike morphology in wheat. Conditional QTL analysis revealed that QTL for SCN were mainly affected by SL. Moreover, a Kompetitive Allele Specific PCR (KASP) marker tightly linked to stable major QTL QPh.cas-5A.3 was developed and verified using the PG-RIL population and a natural population. CONCLUSIONS: Twenty-one stable QTL related to PH, SL, and SCN were identified. These stable QTL and the user-friendly marker KASP8750 will facilitate future studies involving positional cloning and marker-assisted selection in breeding.

Characterization of a new splicing variant of powdery mildew resistance gene Pm4 in synthetic hexaploid wheat YAV249.

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is a destructive fungal disease of wheat throughout the world. Utilization of effective powdery mildew resistance genes and cultivars is considered as the most economic, efficient, and environmental-friendly method to control this disease. Synthetic hexaploid wheat (SHW), which was developed through hybridization of diploid Aegilops and tetraploid wheat, is a valuable genetic resource for resistance to powdery mildew. SHW line YAV249 showed high levels of resistance to powdery mildew at both the seedling and adult stages. Genetic analysis indicated that the resistance was controlled by a single dominant gene, temporarily designated PmYAV. Bulked segregant analysis with wheat 660K single nucleotide polymorphism (SNP) array scanning and marker analysis showed that PmYAV was located on chromosome 2AL and flanked by markers Xgdm93 and Xwgrc763, respectively, with genetic distances of 0.8 cM and 1.2 cM corresponding to a physic interval of 1.89 Mb on the Chinese Spring reference genome sequence v1.0. Sequence alignment analysis demonstrated that the sequence of PmYAV was consistent with that of Pm4a but generated an extra splicing event. When inoculated with different Bgt isolates, PmYAV showed a significantly different spectrum from Pm4a, hence it might be a new resistant resource for improvement of powdery mildew resistance. The flanked markers GDM93 and WGRC763, and the co-segregated markers BCD1231 and JS717/JS718 were confirmed to be easily performed in marker-assisted selection (MAS) of PmYAV. Using MAS strategy, PmYAV was transferred into the commercial cultivar Kenong 199 (KN199) and a wheat line YK13 was derived at generation BC3F3 from the population of YAV249/4*KN199 due to its excellent agronomic traits and resistance to powdery mildew. In conclusion, an alternative splicing variant of Pm4 was identified in this study, which informed the regulation of Pm4 gene function.

PM2b, a CC-NBS-LRR protein, interacts with TaWRKY76-D to regulate powdery mildew resistance in common wheat.

Powdery mildew caused by Blumeria graminis f. sp. tritici (Bgt) is a destructive disease of wheat throughout the world. Host resistance is considered the most sustainable way to control this disease. Powdery mildew resistance gene Pm2b was mapped to the same genetic interval with Pm2a and PmCH1357 cloned previously, but showed different resistance spectra from them, indicating that they might be caused by different resistance genes or alleles. In this study, Pm2b was delimited to a 1.64 Mb physical interval using a large segregating population containing 4,354 F2:3 families of resistant parent KM2939 and susceptible cultivar Shimai 15. In this interval, TraesCS5D03G0111700 encoding the coiled-coil nucleotide-binding site leucine-rich repeat protein (CC-NBS-LRR) was determined as the candidate gene of Pm2b. Silencing by barley stripe mosaic virus-induced gene silencing (BSMV-VIGS) technology and two independent mutants analysis in KM2939 confirmed the candidate gene TraesCS5D03G0111700 was Pm2b. The sequence of Pm2b was consistent with Pm2a/PmCH1357. Subcellular localization showed Pm2b was located on the cell nucleus and plasma membrane. Pm2b had the highest expression level in leaves and was rapidly up-regulated after inoculating with Bgt isolate E09. The yeast two-hybrid (Y2H) and luciferase complementation imaging assays (LCI) showed that PM2b could self-associate through the NB domain. Notably, we identified PM2b interacting with the transcription factor TaWRKY76-D, which depended on the NB domain of PM2b and WRKY domain of TaWRKY76-D. TaWRKY76-D negatively regulated the resistance to powdery mildew in wheat. The specific KASP marker K529 could take the advantage of high-throughput and high-efficiency for detecting Pm2b and be useful in molecular marker assisted-selection breeding. In conclusion, cloning and disease resistance mechanism analysis of Pm2b provided an example to emphasize a need of the molecular isolation of resistance genes, which has implications in marker assisted wheat breeding.

Identification of the major QTL QPm.cas-7D for adult plant resistance to wheat powdery mildew.

Developing effective and durable host plant resistance is crucial for controlling powdery mildew, a devastating disease caused by Blumeria graminis f. sp. tritici (Bgt). In the present study, we dissected the genetic basis of the adult plant resistance to powdery mildew using a recombinant inbred line (RIL) composed of 176 F9 RILs population derived from a cross between PuBing 3228 (P3228) and susceptible cultivar Gao 8901. P3228 exhibits stable adult-plant resistance to powdery mildew in the field over consecutive years. We identified two QTLs on chromosomes 7DS (QPm.cas-7D) and 1AL (QPm.cas-1A) contributed by P3228, and one QTL on 3DS (QPm.cas-3D) contributed by Gao 8901, which could explain 65.44%, 3.45%, and 2.18% of the phenotypic variances, respectively. By analyzing the annotated genes in the 1.168 Mb physical interval of the major QTL QPm.cas-7D, we locked a previously cloned adult-plant resistance gene Pm38 that was most probably the candidate gene of QPm.cas-7D. Sequence alignment analysis revealed that the candidate gene of QPm.cas-7D in P3228 was identical to the reported Pm38 sequence. Two haplotypes QPm-7D-R and QPm-7D-S were identified in the whole Pm38 genomic regions between P3228 and Gao 8901. To apply QPm.cas-7D in wheat breeding, we developed a kompetitive allele-specific PCR (KASP) marker Kasp5249 that is closely linked with these haplotypes. It is worth mentioning that the QPm-7D-R haplotype significantly decreased TKW and underwent negative selection for higher yields in China wheat breeding. In this study, we identified a major QTL QPm.cas-7D and revealed the relationship between its resistance and yield, which could be beneficial for further applications in wheat disease resistance and high-yield breeding.

Molecular Cytogenetic Identification of a New Wheat-Rye 6R Addition Line and Physical Localization of Its Powdery Mildew Resistance Gene.

Rye (Secale cereale L.), a naturally cross-pollinating relative of wheat, is a tertiary gene donor and of substantial value in wheat improvement. Wheat powdery mildew is caused by Blumeria graminis f. sp. tritici (Bgt), which seriously affects yield and quality worldwide. Identifying and transferring new, effective resistance genes against powdery mildew from rye is important for wheat breeding. The current study developed a wheat-rye line YT2 resistant to powdery mildew by crossing, backcrossing, and self-pollination for multiple generations between octoploid triticale 09R2-100 and common wheat cultivar Shixin 616. YT2 was confirmed to be a 6R disomic addition and T1RS⋅1BL translocation line by genomic in situ hybridization (GISH), multicolor fluorescence in situ hybridization (mc-FISH), multicolor-GISH (mc-GISH), and molecular marker analyses. Disease responses to different Bgt isolates and genetic analysis showed that the powdery mildew resistance gene of YT2 was derived from the rye chromosome 6R of 09R2-100, which differed from the previously reported Pm genes from rye including Pm20 on 6RL. Resistance phenotype of different translocation lines and deletion lines derived from YT2 combined with newly developed 6RL-specific markers analysis suggested that the powdery mildew resistance gene of YT2 was localized to the region in chromosome 6RL: 890.09-967.51 Mb and flanked by markers XM189 and X4M19, corresponding to the reference genome of Weining rye. Therefore, YT2 could be used as a promising bridging parent for wheat disease resistance improvement.

A Novel Wheat-Rye 2R (2D) Disomic Substitution Line Pyramids Two Types of Resistance to Powdery Mildew.

Powdery mildew, caused by Blumeria graminis f. sp. tritici, is a devastating disease of wheat that seriously affects yield and quality worldwide. Because of the extensive growth of wheat cultivars with homogeneous genetic background, exploring novel resistant resources from wheat relatives has become important for increasing the genetic diversity of wheat. Rye (Secale cereale) is a wheat relative possessing abundant resistance genes because of its high variation. Wheat line AL69, resistant to powdery mildew, was developed by crossing, backcrossing, and self-pollination for multiple generations between hexaploid triticale Zhongsi 237 and common wheat cultivar Zimai 17. Through genomic in situ hybridization (GISH) and multicolor fluorescence in situ hybridization (FISH), nondenaturing FISH, multicolor GISH, and selection with specific molecular markers, AL69 was determined to be a wheat-rye 2R (2D) disomic substitution line. Testing with different B. graminis f. sp. tritici isolates and genetic analysis showed that the all-stage resistance (also called seedling resistance) of AL69 was conferred by the cataloged powdery mildew resistance gene Pm4b derived from Zimai 17, and its adult-plant resistance was derived from the alien chromosome 2R of Zhongsi 237, which was found to be different from the previously reported rye-derived Pm genes, including Pm7 on 2RL. In addition, AL69 showed improved spike number per plant, spike length, fertile spikelet number per spike, kernel number per spike, and grain yield per plant compared with its wheat parent Zimai 17. An elite line S251 combining powdery mildew resistance with excellent agronomic performance was selected from the progenies of AL69 and wheat cultivar Jimai 22. Therefore, AL69 has two types of resistance genes to powdery mildew and improved agronomic traits through pyramiding and thus can be used as a promising genetic stock for wheat breeding.

Identification of Resistant Germplasm and Detection of Genes for Resistance to Powdery Mildew and Leaf Rust from 2,978 Wheat Accessions.

Powdery mildew and leaf rust, caused by Blumeria graminis f. sp. tritici and Puccinia triticina, respectively, are widespread diseases of wheat worldwide. The use of resistant cultivars is considered the most economical, environment-friendly, and effective method to control these diseases. In the present study, a collection of 2,978 wheat accessions consisting of 1,394 advanced breeding lines, 1,078 Chinese cultivars, 291 introduced cultivars, 132 lines containing alien chromosomes, and 83 landraces was tested for reactions to powdery mildew and leaf rust. The results indicated that 659 wheat accessions (22.1%) were highly resistant to a widely prevalent B. graminis f. sp. tritici isolate, E09, at the seedling stage, and 390 were consistently resistant to the mixture of B. graminis f. sp. tritici isolates at the adult plant stage. Meanwhile, 63 accessions (2.1%) were highly resistant to leaf rust at the adult plant stage, of which 54 were resistant to a predominant and highly virulent P. triticina race, THTT, at the seedling stage. Notably, 17 accessions were resistant to both powdery mildew and leaf rust. To detect known genes for resistance to powdery mildew and leaf rust, these accessions were tested with gene-specific or tightly linked markers for seven powdery mildew genes (Pm genes; Pm2, Pm4, Pm5, Pm6, Pm8, Pm21, and Pm24) and 10 Lr genes (Lr1, Lr9, Lr10, Lr19, Lr20, Lr24, Lr26, Lr34, Lr37, and Lr46). Of the 659 powdery mildew-resistant accessions, 328 might carry single Pm genes and 191 carry combined Pm genes. Pm2 was detected at the highest frequency of 59.6%, followed by Pm8, Pm6, Pm21, Pm4, and Pm5, whereas Pm24 was not detected. In addition, 139 accessions might contain unknown Pm genes different from those tested in this study. In the 63 accessions resistant to leaf rust, four leaf rust genes (Lr genes; Lr1, Lr10, Lr26, and Lr34) were detected in 41 accessions singly or in combination, whereas six genes (Lr9, Lr19, Lr20, Lr24, Lr37, and Lr46) were not detected. Twenty-two accessions might contain unknown Lr genes different from those tested in this study. This study not only provided important information for rationally distributing resistance genes in wheat breeding programs, but also identified resistant germplasm that might have novel genes to enrich the diversity of resistance sources.

Identification and validation of quantitative trait loci for kernel traits in common wheat (Triticum aestivum L.).

BACKGROUND: Kernel weight and morphology are important traits affecting cereal yields and quality. Dissecting the genetic basis of thousand kernel weight (TKW) and its related traits is an effective method to improve wheat yield. RESULTS: In this study, we performed quantitative trait loci (QTL) analysis using recombinant inbred lines derived from the cross 'PuBing3228 × Gao8901' (PG-RIL) to dissect the genetic basis of kernel traits. A total of 17 stable QTLs related to kernel traits were identified, notably, two stable QTLs QTkw.cas-1A.2 and QTkw.cas-4A explained the largest portion of the phenotypic variance for TKW and kernel length (KL), and the other two stable QTLs QTkw.cas-6A.1 and QTkw.cas-7D.2 contributed more effects on kernel width (KW). Conditional QTL analysis revealed that the stable QTLs for TKW were mainly affected by KW. The QTLs QTkw.cas-7D.2 and QKw.cas-7D.1 associated with TKW and KW were delimited to the physical interval of approximately 3.82 Mb harboring 47 candidate genes. Among them, the candidate gene TaFT-D1 had a 1 bp insertions/deletion (InDel) within the third exon, which might be the reason for diversity in TKW and KW between the two parents. A Kompetitive Allele-Specific PCR (KASP) marker of TaFT-D1 allele was developed and verified by PG-RIL and a natural population consisted of 141 cultivar/lines. It was found that the favorable TaFT-D1 (G)-allele has been positively selected during Chinese wheat breeding. Thus, these results can be used for further positional cloning and marker-assisted selection in wheat breeding programs. CONCLUSIONS: Seventeen stable QTLs related to kernel traits were identified. The stable QTLs for thousand kernel weight were mainly affected by kernel width. TaFT-D1 could be the candidate gene for QTLs QTkw.cas-7D.2 and QKw.cas-7D.1.

Identification of an Elite Wheat-Rye T1RS·1BL Translocation Line Conferring High Resistance to Powdery Mildew and Stripe Rust.

Wheat-rye T1RS·1BL translocations have been widely used worldwide in wheat production for multiple disease resistance and superior yield traits. However, many T1RS·1BL translocations have successively lost their resistance to pathogens due to the coevolution of pathogen virulence with host resistance. Because of the extensive variation in rye (Secale cereale L.) as a naturally cross-pollinating relative of wheat, it still has promise to widen the variation of 1RS and to fully realize its application value in wheat improvement. In the present study, the wheat-rye breeding line R2207 was characterized by comprehensive analyses using genomic in situ hybridization (GISH), multicolor fluorescence in situ hybridization with multiple probes, multicolor GISH, and molecular marker analysis, and then was proven to be a cytogenetically stable wheat-rye T1RS·1BL translocation line. Based on the disease responses to different isolates of powdery mildew and genetic analysis, R2207 appears to possess a novel variation for resistance, which was confirmed to be located on the rye chromosome arm 1RS. Line R2207 also exhibited high levels of resistance to stripe rust at both seedling and adult stages, as well as enhanced agronomic performance, so it has been transferred into a large number of commercial cultivars using an efficient 1RS-specific kompetitive allele specific PCR marker for marker-assisted selection.

Scale development and utilization of universal PCR-based and high-throughput KASP markers specific for chromosome arms of rye (Secale cereale L.).

BACKGROUND: Rye (Secale cereale L., 2n = 2x = 14, RR), a relative of common wheat, is a large gene resource pool for wheat improvement. Accurate and convenient identification of the rye chromatin in wheat background will facilitate the transfer and utilization of elite genes derived from rye in wheat breeding. RESULTS: In the present study, five rye cultivars including Imperial, German White, Jingzhouheimai, Baili and Guyuan were sequenced by specific-locus amplified fragment sequencing (SLAF-seq) to develop large-scale rye-specific markers. Based on SLAF-seq and bioinformatics analyses, a total of 404 universal PCR-based and a whole set of Kompetitive allele-specific PCR (KASP) markers specific for the 14 individual rye chromosome arms were developed and validated. Additionally, two KASP markers specific for 1RS and 2RL were successfully applied in the detection of 1RS translocations in a natural population and 2RL chromosome arms in wheat-rye derived progenies that conferred adult resistance to powdery mildew. CONCLUSION: The 404 PCR-based markers and 14 KASP markers specific for the 14 individual rye chromosome arms developed in this study can enrich the marker densities for gene mapping and accelerate the utilization of rye-derived genes in wheat improvement. Especially, the KASP markers achieved high-throughput and accurate detection of rye chromatin in wheat background, thus can be efficiently used in marker-assisted selection (MAS). Besides, the strategy of rye-specific PCR-based markers converting into KASP markers was high-efficient and low-cost, which will facilitate the tracing of alien genes, and can also be referred for other wheat relatives.

TaDA1, a conserved negative regulator of kernel size, has an additive effect with TaGW2 in common wheat (Triticum aestivum L.).

Kernel size is an important trait determining cereal yields. In this study, we cloned and characterized TaDA1, a conserved negative regulator of kernel size in wheat (Triticum aestivum). The overexpression of TaDA1 decreased the size and weight of wheat kernels, while its down-regulation using RNA interference (RNAi) had the opposite effect. Three TaDA1-A haplotypes were identified in Chinese wheat core collections, and a haplotype association analysis showed that TaDA1-A-HapI was significantly correlated with the production of larger kernels and higher kernel weights in modern Chinese cultivars. The haplotype effect resulted from a difference in TaDA1-A expression levels between genotypes, with TaDA1-A-HapI resulting in lower TaDA1-A expression levels. This favourable haplotype was found having been positively selected during wheat breeding over the last century. Furthermore, we demonstrated that TaDA1-A physically interacts with TaGW2-B. The additive effects of TaDA1-A and TaGW2-B on kernel weight were confirmed not only by the phenotypic enhancement arising from the simultaneous down-regulation of TaDA1 and TaGW2 expression, but also by the combinational haplotype effects estimated from multi-environment field data from 348 wheat cultivars. A comparative proteome analysis of developing transgenic and wild-type grains indicated that TaDA1 and TaGW2 are involved in partially overlapping but relatively independent protein regulatory networks. Thus, we have identified an important gene controlling kernel size in wheat and determined its interaction with other genes regulating kernel weight, which could have beneficial applications in wheat breeding.

Development of Novel Wheat-Rye Chromosome 4R Translocations and Assignment of Their Powdery Mildew Resistance.

Rye (Secale cereale L.) is an important gene donor for wheat improvement because of its many valuable traits, especially disease resistance. Development of novel wheat-rye translocations with disease resistance can contribute to transferring resistance into common wheat. In a previous study, a wheat-rye T4BL·4RL and T7AS·4RS translocation line (WR41-1) was developed by distant hybridization, and it was speculated that its resistance to powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), was derived from rye based on pedigree analysis. To make accurate use of chromosome 4R in wheat improvement, a set of new 4R translocations involving different arm translocations (e.g., 4RS monosomic, 4RL monosomic, 4RL disomic, 4RS monosomic plus 4RL monosomic, 4RS monosomic plus 4RL disomic, and 4RS disomic plus 4RL disomic translocations) was developed from crosses with common wheat. Those translocations were characterized by genomic in situ hybridization and expressed sequence tag simple sequence repeat marker analysis. To confirm the source of powdery mildew resistance, the translocation plants were tested against Bgt isolate E09. The results indicated that all translocations with 4RL were resistant at all tested growth stages, whereas those with only 4RS translocation or no alien translocation were susceptible. This further indicated that the powdery mildew resistance of WR41-1 was derived from the alien chromosome arm 4RL. To effectively use 4RL resistance in wheat improvement, two competitive allele-specific PCR markers specific for chromosome arm 4RL were developed to detect the alien chromosome in the wheat genome. These new translocation lines with diagnostic markers can efficiently serve as important bridges for wheat improvement.

Identification of a New Powdery Mildew Resistance Gene pmDHT at or Closely Linked to the Pm5 Locus in the Chinese Wheat Landrace Dahongtou.

Chinese wheat landrace Dahongtou was resistant to 35 of 38 tested Chinese isolates of Blumeria graminis f. sp. tritici at the seedling stage. Genetic analysis of the F2 populations and their derived F2:3 families of crosses of Dahongtou with the susceptible varieties Mingxian 169 and Huixianhong indicated that the resistance of Dahongtou to B. graminis f. sp. tritici isolate E09 was conferred by a single recessive gene, tentatively designated as pmDHT. The gene was mapped to chromosome arm 7BL and flanked by markers Xwmc526/XBE443877 and Xgwm611/Xwmc511 at genetic distances of 0.8 and 0.3 cM, respectively. The chromosomal position of pmDHT was similar to the multi-allelic Pm5 locus on 7BL. Allelism tests with crosses of Dahongtou with Fuzhuang 30 (Pm5e) and Xiaobaidong (mlxbd) indicated that pmDHT was allelic to both Pm5e and mlxbd. However, pmDHT showed a different pattern of resistance to the 38 B. graminis f. sp. tritici isolates compared with wheat lines with Pm5a, Pm5b, Pm5e, mlxbd, and PmHYM and also differed from PmSGA. Thus, pmDHT was identified most likely as a new allele or at least a closely linked gene of the Pm5 locus. This gene can be transferred into susceptible wheat cultivars/lines and pyramided with other resistance genes through marker-assisted selection to improve powdery mildew resistance.