Suoping Li

Key message A candidate gene TaSP1 related to spike shape was cloned, and the gene-specific marker was developed to efficiently track the superior haplotype in common wheat. Abstract Spike shape, an important factor that affects wheat grain yield, is mainly defined by spike length (SPL), spikelet number (SPN), and compactness. Zhoumai32 mutant 1160 (ZM1160), a mutant obtained from ethyl methane sulfonate (EMS) treatment of hexaploid wheat variety Zhoumai32, was used to identify and clone the candidate gene that conditioned the spike shape. Genetic analysis of an F₂ population derived from a cross of ZM1160 and Bainong207 suggested that the compact spike shape in ZM1160 was controlled by a single recessive gene, and therefore, the mutated gene was designated as Tasp1. With polymorphic markers identified through bulked segregant analysis (BSA), the gene was mapped to a 2.65-cM interval flanked by markers YZU0852 and MIS46239 on chromosome 7D, corresponding to a 0.42-Mb physical interval of Chinese spring (CS) reference sequences (RefSeq v1.0). To fine map TaSP1, 15 and seven recombinants were, respectively, screened from 1599 and 1903 F₃ plants derived from the heterozygous F₂ plants. Finally, TaSP1 was delimited to a 21.9 Kb (4,870,562 to 4,892,493 bp) Xmis48123-Xmis48104 interval. Only one high-confidence gene TraesCS7D02G010200 was annotated in this region, which encodes an unknown protein with a putative vWA domain. Quantitative reverse transcription PCR (qRT-PCR) analysis showed that TraesCS7D02G010200 was mainly expressed in the spike. Haplotype analysis of 655 wheat cultivars using the candidate gene-specific marker Xg010200p2 identified a superior haplotype TaSP1b with longer spike and more spikelet number. TaSP1 is beneficial to the improvement in wheat spike shape.

Key message The powdery mildew resistance gene Pm58 was traced to a 141.3-kb interval with the co-segregating marker Xkasp68500 in wheat breeding. Abstract Pm58 is a powdery mildew resistance gene identified in Aegilops tauschii accession TA1662 and effective in a common wheat background. To finely map Pm58, an F₂ population of 676 plants derived from the cross T093 × TA1662 was used for recombinant screening. We obtained 13 recombinants that occurred between the flanking markers Xhnu670 and Xhnu186. Genotyping and phenotyping these recombinant F_2:3 families delimited Pm58 to a 0.22-cM interval (Xsts20220–Xkasp61553) on chromosome arm 2DS. The region carrying the Pm58 locus was approximately 141.3-kb, which contained eight annotated genes according to the reference genome sequence of Ae. tauschii AL8/78. Haplotype analysis of 178 Ae. tauschii accessions using the candidate gene-specific markers identified a disease resistance gene AET2Gv20068500 as a candidate for Pm58. Comparative mapping of the Pm58-containing interval revealed two presence/absence variations (PAVs) between AL8/78 and common wheat Chinese Spring. PAV-1 resides in the 3′-end of AET2Gv20068500. The majority of 158 common wheat cultivars (84.8%) displayed the absence of a 14.1-kb fragment in the PAV-1 region, which was confirmed by aligning the targeted genome sequences of the other sequenced Ae. tauschii accessions and common wheat cultivars. A co-segregating marker Xkasp68500 developed from AET2Gv20068500 can distinguish TA1662 from all randomly selected common wheat cultivars and will be instrumental for tracking Pm58 in breeding programs.

High molecular weight glutenin subunits (HMW-GS) and Low molecular weight glutenin subunits (LMW-GS) in mature grains play important roles in the formation of glutenin macropolymer and gluten quality. To characterize the expressed glutenin genes of the bread wheat variety Xinmai 26 during seed development, in this study, we measured the dough rheological properties of mature grains through farinograph and gluten testing system, and revealed its strong gluten quality. The compositions of HMW-GS and LMW-GS were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Further, a total of 18 full-length transcripts were obtained by third-generation RNA sequencing, including 5 transcripts of HMW-GS genes and 13 transcripts of LMW-GS genes (8 intact genes and 5 pseudogenes). The deduced protein structures of transcript sequences exhibit the typical HMW-GS and LMW-GS structural characteristics. Moreover, a specific functional marker was developed to make better use of the extra cysteine residue of 1Dx5 subunit. This study provides an efficient method to accurately identify glutenin genes in bread wheat through matching full-length transcripts to their spectrum of glutenin, which will be helpful to understand the contributions of glutenin genes to wheat quality.

Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici, is a devastating disease that threatens yield and quality. Host resistance is considered the most effective and preferred means to control this disease. Wheat landrace Duanganmang (DGM) showed high resistance or near immunity to Blumeria graminis f. sp. tritici mixture from Henan Province, China. DGM was crossed with highly susceptible Chinese wheat landrace Huixianhong (HXH) and cultivar ‘Shimai 15’ (SM15) to produce genetic populations. The resistance of DGM to Blumeria graminis f. sp. tritici isolate E09 was shown to be controlled by a single dominant Mendelian factor, tentatively designated PmDGM. Marker analysis and 55K single nucleotide polymorphism (SNP) array scanning showed that this gene was positioned in the Pm5 interval (2.4 cM or 1.61 Mb) flanked by Xhenu099 and Xmp1158 in the Chinese Spring reference genome. Homology-based cloning and sequence analysis demonstrated that DGM has the identical NLR gene (Pm5e) and RXL gene reported in Fuzhuang 30 (FZ30), conferring and modifying powdery mildew resistance, respectively. However, based on the different reaction patterns to the Blumeria graminis f. sp. tritici isolate B15 between DGM and FZ30, the authors speculate that DGM may have two tightly linked genes that could not be separated in the current mapping population, one of which is PmDGM and the other being Pm5e. Hence, this study provides a valuable resistance resource for improvement of powdery mildew resistance.

Powdery mildew, caused by fungal pathogen Blumeria graminis f. sp. tritici, is an agronomically important and widespread wheat disease causing severe yield losses. Deployment of broad‐spectrum disease resistance genes is the preferred strategy to prevent this pathogen. Chinese wheat landrace Honghuaxiaomai (HHXM) was resistant to all 23 tested B. graminis f. sp. tritici isolates at the seedling stage. The F₁, F₂, and F_2:3 progenies derived from the cross HHXM × Yangmai 158 were used in this study, and genetic analysis revealed that a single dominant gene, designated PmHHXM, conferred resistance to B. graminis f. sp. tritici isolate E09. Bulked segregant analysis and molecular mapping initially located PmHHXM to the distal region of chromosome 4AL. To fine map PmHHXM, we identified two critical recombinants from 592 F₂ plants and delimited PmHHXM to a 0.18-cM Xkasp475200 to Xhnu552 interval covering 1.77 Mb, in which a number of disease resistance–related gene clusters were annotated. Comparative mapping of this interval revealed a perturbed synteny among Triticeae species. This study reports the new powdery mildew resistance gene PmHHXM, which seems different from three known quantitative trait loci/genes identified on chromosome 4AL and has significant values for further genetic improvement. Analysis of the polymorphisms of 13 cosegregating markers between HHXM and 170 modern wheat cultivars indicates that Xhnu227 and Xsts478700 developed here are ideal for marker-assisted introgression of this locus in wheat breeding.

Wheat pre-harvest sprouting (PHS) causes serious losses in wheat yield. In this study, precise mapping was carried out in the chromosome segment substitution lines (CSSL) F₂ population generated by a direct cross of Zhoumai 18 (PHS-sensitive) and Aegilops tauschii accession T093 (highly PHS-resistant). Three Ae. tauschii-derived quantitative trait loci (QTLs), QDor.3D.1, QDor.3D.2, and QDor.3D.3, were detected on chromosome 3DL using four simple sequence repeats (SSR) markers and 10 developed Kompetitive allele-specific PCR (KASP) markers. Alongside these QTL results, the RNA-Seq and qRT-PCR analysis revealed expression levels of TraesCS3D01G466100 in the QDor.3D.2 region that were significantly higher in CSSLs 495 than in Zhoumai 18 during the seed imbibition treatment. The cDNA sequencing results of TraesCS3D01G466100 showed two single nucleotide polymorphisms (SNPs), resulting in two changed amino acid substitutions between Zhoumai 18 and line 495, and the 148 nt amino acid substitution of TraesCS3D01G466100, derived from Ae. tauschii T093, which may play an important role in the functioning of ubiquitin ligase enzymes 3 (E3) according to the homology protein analysis, which could lead to differential PHS-resistance phenotypes. Taken together, our results may foster a better understanding of the mechanism of PHS resistance and are potentially valuable for marker-assisted selection in practical wheat breeding efforts.

As the diploid progenitor of common wheat, Aegilops tauschii is considered to be a valuable resistance source to various biotic and abiotic stresses. However, little has been reported concerning the molecular mechanism of drought tolerance in Ae. tauschii. In this work, the drought tolerance of 155 Ae. tauschii accessions was firstly screened on the basis of their coleoptile lengths under simulated drought stress. Subsequently, two accessions (XJ002 and XJ098) with contrasting coleoptile lengths were selected and intensively analyzed on rate of water loss (RWL) as well as physiological characters, confirming the difference in drought tolerance at the seedling stage. Further, RNA-seq was utilized for global transcriptome profiling of the two accessions seedling leaves under drought stress conditions. A total of 6969 differentially expressed genes (DEGs) associated with drought tolerance were identified, and their functional annotations demonstrated that the stress response was mediated by pathways involving alpha-linolenic acid metabolism, starch and sucrose metabolism, peroxisome, mitogen-activated protein kinase (MAPK) signaling, carbon fixation in photosynthetic organisms, and glycerophospholipid metabolism. In addition, DEGs with obvious differences between the two accessions were intensively analyzed, indicating that the expression level of DEGs was basically in alignment with the physiological changes of Ae. tauschii under drought stress. The results not only shed fundamental light on the regulatory process of drought tolerance in Ae. tauschii, but also provide a new gene resource for improving the drought tolerance of common wheat.

Nucleotide‐binding (NB) leucine‐rich repeat (LRR) receptors (NLRs) provide resistance against several plant pathogens. We previously cloned the wheat powdery mildew resistance gene Pm21, which encodes a coiled‐coil (CC) NLR that confers broad‐spectrum resistance against Blumeria graminis f. sp. tritici. Here, we report comprehensive biochemical and functional analyses of Pm21 CC domain in Nicotiana benthamiana. Transient overexpression assay suggested that only the extended CC (eCC, amino acid residues 1–159) domain has cell‐death‐inducing activity, whereas the CC‐containing truncations, including CC‐NB and CC‐NB‐LRR, do not induce cell‐death responses. Coimmunoprecipitation (Co‐IP) assay showed that the eCC domain self‐associates and interacts with the NB and LRR domains in planta. These results imply that the activity of the eCC domain is inhibited by the intramolecular interactions of different domains in the absence of pathogens. We found that the LRR domain plays a crucial role in D491V‐mediated full‐length (FL) Pm21 autoactivation. Some mutations in the CC domain leading to the loss of Pm21 resistance to powdery mildew impaired the CC activity of cell‐death induction. Two mutations (R73Q and E80K) interfered with D491V‐mediated Pm21 autoactivation without affecting the cell‐death‐inducing activity of the eCC domain. Notably, some susceptible mutants harbouring mutations in the CC domain still exhibited cell‐death‐inducing activity. Taken together, these results implicate the CC domain of Pm21 in cell‐death signalling and disease‐resistance signalling, which are potentially independent of each other.

The D genome progenitor of bread wheat, Aegilops tauschii Cosson (DD, 2n = 2x = 14), which is naturally distributed in Central Eurasia, ranging from northern Syria and Turkey to western China, is considered a potential genetic resource for improving bread wheat. In this study, the chloroplast (cp) genomes of 17 Ae. tauschii accessions were reconstructed. The cp genome sizes ranged from 135,551 bp to 136,009 bp and contained a typical quadripartite structure of angiosperms. Within these genomes, we identified a total of 124 functional genes, including 82 protein-coding genes, 34 transfer RNA genes and eight ribosomal RNA genes, with 17 duplicated genes in the IRs. Although the comparative analysis revealed that the genomic structure (gene order, gene number and IR/SC boundary regions) is conserved, a few variant loci were detected, predominantly in the non-coding regions (intergenic spacer regions). The phylogenetic relationships determined based on the complete genome sequences were consistent with the hypothesis that Ae. tauschii populations in the Yellow River region of China originated in South Asia not Xinjiang province or Iran, which could contribute to more effective utilization of wild germplasm resources. Furthermore, we confirmed that Ae. tauschii was derived from monophyletic speciation rather than hybrid speciation at the cp genome level. We also identified four variable genomic regions, rpl32-trnL-UAG, ccsA-ndhD, rbcL-psaI and rps18-rpl20, showing high levels of nucleotide polymorphisms, which may accordingly prove useful as cpDNA markers in studying the intraspecific genetic structure and diversity of Ae. tauschii.