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Article

Genetic Dissection of the Powdery Mildew Resistance in a Cultivated Emmer Wheat Accession

by
Ruishan Liu
1,†,
Yuli Jin
1,†,
Ningning Yu
1,
Hongxing Xu
2,
Xusheng Sun
3,
Jiangchun Wang
4,5,
Xueqing Liu
4,
Jiadong Zhang
1,
Jiatong Li
1,
Yaoxue Li
1 and
Pengtao Ma
1,*
1
Yantai Key Laboratory of Characteristic Agricultural Biological Resources Conservation and Germplasm Innovative Utilization, College of Life Sciences, Yantai University, Yantai 264005, China
2
School of Life Sciences, Henan University, Kaifeng 475004, China
3
Yantai Agricultural Technology Extension Center, Yantai 264001, China
4
Yantai Academy of Agricultural Sciences, Yantai 265500, China
5
Shandong Zhongnong Tiantai Seed Industry Co., Ltd., Linyi 273300, China
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Agronomy 2025, 15(4), 980; https://doi.org/10.3390/agronomy15040980
Submission received: 11 March 2025 / Revised: 15 April 2025 / Accepted: 16 April 2025 / Published: 18 April 2025
(This article belongs to the Special Issue Mechanism and Sustainable Control of Crop Diseases)
Browse Figures
Versions Notes

Abstract

:
Blumeria graminis f. sp. tritici (Bgt), the causal agent of wheat powdery mildew, poses a significant threat to global wheat production. In this study, we identified and characterized a broad-spectrum powdery mildew resistance gene, PmL709, in a resistant cultivated emmer wheat (Triticum dicoccum) accession: L709. Using bulked segregant RNA sequencing (BSR-Seq) analysis and molecular markers, PmL709 was mapped to a 1.7 cM interval on chromosome arm 2BS, flanked by markers Xdw05/YTU95-04/YTU95-06/YTU95-08/Xdw10/Xdw11 and YTU692B-094, corresponding to a 21.82–25.94 Mb physical interval (cv. Svevo), using the segregated population crossed by L709 and a susceptible durum wheat cultivar, Langdon. Referring to the origin, the resistance spectra, and the physical position with known resistance genes on chromosome arm 2BS, PmL709 was likely to be an allele of Pm68. Transcriptomic analysis revealed 3923 differentially expressed genes (DEGs) between resistant and susceptible bulks, enriched in pathways such as phenylpropanoid biosynthesis, MAPK signaling, and plant–pathogen interactions. qRT-PCR validated the differential expression of nine candidate genes within the PmL709 interval, highlighting their potential roles in disease resistance. The flanking markers could accurately trace the presence of PmL709 from resistant accession L709 in a survey of 46 susceptible wheat accessions. These findings provide valuable insights into the genetic and molecular mechanisms of powdery mildew resistance in wheat and offer practical tools for marker-assisted breeding to develop resistant cultivars.

1. Introduction

The widespread cultivation of common wheat (Triticum aestivum L.) has positioned it as a key contributor to maintaining stable food supplies across the world [1]. Despite its global cultivation, wheat remains vulnerable to multiple phytopathological challenges, notably infections from Blumeria graminis f. sp. tritici (Bgt) [2]. Wheat powdery mildew, triggered by the obligate fungal pathogen Bgt, severely jeopardizes crop yields, with reported losses reaching 50% in high-infection scenarios [3,4,5]. Beyond conventional chemical control practices, the employment of resistant wheat cultivars represents a key strategy for managing powdery mildew, serving as the most cost-effective solution in integrated disease management systems.
Up to the present, over 140 powdery mildew resistance genes and/or alleles have been characterized and localized within wheat and its wild relative species, including 71 officially designated Pm genes (Pm1-–Pm71) and dozens of temporarily named genes [6,7]. Only a small number of these genes have been cloned. Among them, genes including Pm1a, Pm2a, Pm3b/Pm8, Pm5e, Pm12, Pm21, Pm41, Pm60/MlIW172, and Pm69 are characterized as encoding nucleotide-binding leucine-rich repeat (NLR) proteins, which serve as central mediators in plant defense responses [8,9,10,11]. Functional genomic studies have revealed distinct protein architectures in resistance genes: Pm24 encodes a tandem kinase protein [12], while Pm4 generates a chimeric polypeptide characterized by a pseudokinase domain (serine/threonine specificity), multiple C2 domains, and a transmembrane structural motif. Significantly, two cloned resistance genes, Pm38 (also known as Lr34/Yr18/Sr57 encoding an ABC transporter) and Pm46 (Lr67/Yr46/Sr55 encoding a hexose transporter), have been identified to confer broad-spectrum disease resistance against both powdery mildew and rust in wheat [13,14].
The plant immune system comprises two core branches. The first branch employs transmembrane pattern recognition receptors to detect slowly evolving microbe/pathogen-associated molecular patterns, thereby triggering pattern-triggered immunity [15,16]. The second branch is primarily mediated by intracellular NLR proteins, which recognize pathogen effectors through their polymorphic domains to activate effector-triggered immunity (ETI) [17,18]. NLR proteins exhibit remarkable functional specificity: their NB domain binds nucleotides and transduces signaling, while the LRR domain facilitates effector recognition. This modular architecture enables NLRs to generate immune responses against effectors from diverse pathogen lineages [19]. Notably, the ETI pathway demonstrates significantly higher defense efficiency against obligate biotrophic pathogens compared to necrotrophic pathogens. In wheat, 12 out of 17 cloned powdery mildew resistance genes encode NLR proteins. For instance, Pm2a directly interacts with the transcription factor TaWRKY76-D via its NB domain and the WRKY domain of TaWRKY76-D, a protein–protein interaction critical for regulating disease resistance signaling [18]. Recent studies further revealed that TaWRKY76-D negatively regulates Pm2a expression by binding to its promoter region, uncovering a finely tuned regulatory network within the ETI pathway [20]. These findings provide novel insights into the molecular mechanisms underlying wheat–powdery mildew interactions.
Powdery mildew resistance genes have been ineffective due to the accelerated evolution of causal pathogens. Specifically, in major wheat-producing regions of China, the resistance mediated by genes such as Pm1, Pm2, Pm3, Pm5, and Pm8 has undergone substantial reduction [6,21]. Consequently, the discovery and cloning of novel resistance genes, along with their integration into cultivated wheat germplasms, are essential for diversifying the genetic resources available for wheat disease resistance breeding.
Wheat relatives are valuable sources of resistance genes against powdery mildew. These genes often display greater genetic diversity and elevate resistance against variants of Bgt isolates. Notable examples include Pm12, sourced from Aegilops speltoides Tausch [22], and Pm21, derived from Dasypyrum villosum L. Candargy [23]. The identified genes can serve as invaluable genetic resources for developing durable resistance to powdery mildew through targeted trait introgression [24].
Common wheat, characterized as an allohexaploid, possesses an extensive and intricate genomic architecture that creates substantial obstacles for identifying advantageous genetic components and unraveling disease resistance pathways [25]. To address these genomic complexity challenges, researchers have developed innovative approaches, including bulked segregant RNA sequencing (BSR-Seq) technology. BSR-Seq, an approach integrating bulked segregant analysis with RNA sequencing, has emerged as a powerful tool in genomics research for complex polyploid species. This methodology operates independently of pre-existing expressed gene databases, enabling unbiased profiling of transcriptomic landscapes through direct RNA-seq quantification [26,27]. By circumventing limitations imposed by genome complexity, BSR-Seq simultaneously captures sequence and expression data for nearly all transcripts in specific cell/tissue types at defined developmental stages, rendering it particularly suitable for rapid gene mapping in crops with intricate genomes such as wheat and its wild relatives [24,28,29]. This technology has been instrumental in localizing and cloning wheat powdery mildew resistance genes. Specifically, the application of BSR-Seq on F2-derived resistant/susceptible bulks from Jimai 22/Avocet S identified ten single nucleotide polymorphisms (SNPs) within a 12.09 Mb physical interval on chromosome 2AL. These SNPs were subsequently used to develop molecular markers, facilitating the construction of a genetic linkage map of YrJ22 [30]. Moreover, BSR-Seq has been utilized in the analysis and identification of PmQ in the wheat landrace Qingxinmai [31], the genetic dissection of powdery mildew resistance in wheat line LS5082 [32], the expression profiling of powdery mildew resistance in wild emmer wheat D430 [33], and the transcriptomic analysis of powdery mildew resistance mechanisms in wheat cultivar Jimai 23 [34].
Cultivated emmer wheat (T. dicoccum, 2n = 4x = 28, AABB), the tetraploid ancestor of common wheat, evolved from its wild progenitor approximately 10,000 years ago through early agricultural practices [35]. It has evolved through natural selection, hybridization, and artificial selection [36], harboring abundant genetic resources that enhance its resistance to both biotic and abiotic stresses [37,38]. In our lab, a cultivated emmer wheat accession, L709, exhibited stable and robust resistance against powdery mildew pathogens across multiple years in field evaluations. To better explore and utilize powdery mildew resistance in L709, this study investigated its resistance through the construction of genetic segregation populations, molecular identification, resistance microscopy observation, and BSR-seq analysis. Specifically, we crossed L709 with the susceptible durum wheat cultivar Langdon (LDN) to generate F1, F2, and F2:3 families. Genetic inheritance pattern analysis was conducted using molecular identification and resistance microscopy observation. Subsequently, BSR-seq and linkage mapping were integrated to identify candidate resistance loci. This study aimed to (i) evaluate the resistance of L709 to powdery mildew and determine its genetic inheritance pattern by constructing an L709/LDN genetic segregation population; (ii) identify candidate genomic regions using BSR-seq and genetic linkage mapping and determine potential regulatory genes associated with this resistance through transcriptome sequencing; and (iii) characterize the key regulatory genes underlying powdery mildew resistance via BSR-seq analysis and quantitative real-time PCR (qRT-PCR) validation. Elucidating the molecular mechanisms of resistance-related genes in L709 will not only deepen our understanding of wheat defense responses to powdery mildew but also provide novel genetic resources for developing wheat cultivars with broad-spectrum and durable resistance traits.

2. Materials and Methods

2.1. Plant Materials and Pathogens

Cultivated emmer wheat L709, provided by Professor Xu Hongxing from Henan University, showed high resistance to powdery mildew in the field for consecutive years and was preserved in our lab as a breeding parent against wheat powdery mildew. The susceptible durum wheat cultivar LDN was used as a susceptible parent to cross with L709 to develop segregating populations (F1, F2, and F2:3) for subsequent genetic analysis and gene mapping. Ten F1 seeds derived from the L709 × LDN cross were sown to harvest F2 seeds. A total of 246 F2 seeds were selected for phenotypic evaluation to determine the genetic segregation ratio in F2 generation. To confirm the genotypes of resistant F2 plants and revalidate their phenotypes, the 246 F2 plants were transplanted to the field after scoring, and corresponding F2:3 family seeds were harvested. Each F2:3 family with 20 seeds was subsequently planted to verify the genetic segregation ratio of the F2:3 populations. Mingxian 169, a powdery mildew-susceptible wheat cultivar, served as the susceptible check.
In previous studies, six Pm genes from diversified gene donors, including Pm26 from TTD140 (T. turgidum var. dicoccoides) [39], pm42 from G-303-1M (T. turgidum var. dicoccoides) [40,41], PmXQ-0508 from XQ-0508 (T. aestivum L.) [40], Pm68 from TIR 1796 (T. turgidum L. var. durum Desf) [42], MlIW170 from IW170 (T. turgidum var. dicoccoides) [43], and PmWE99 from WE99 (Thinopyrum intermedium) [44] have been reported on the same chromosome arm, 2BS, with gene PmL709 in L709. To identify the relationship between PmL709 and those genes, these six gene donors with L709 were tested against 27 Bgt isolates collected from different wheat-producing regions in China: E05, E07, E09, E15, E17, E18, E20, E21, E23, E31, E32, F01, F02, F05, F06, F07, F08, F09, F10, F16, F17, F18, F19, F21, F23, F25, and F28. These isolates exhibit distinct biological properties across four key dimensions: pathogenicity, genetic characteristics, geographical distribution, and host specificity [45,46].
To facilitate the transfer of the targeted Pm gene(s) in L709, 46 susceptible wheat genotypes which come from multiple major wheat-producing areas and possess excellent agronomic and yield traits were selected to evaluate the polymorphisms of closely linked markers for marker-assisted selection (MAS).

2.2. Phenotypic Identification of Bgt Isolates

Leaf tissue samples of 30 homozygous resistant plants from 30 F2:3 homozygous resistant families and 30 homozygous susceptible plants from 30 F2:3 homozygous susceptible families were collected 14 days post infection and equally pooled to construct resistant and susceptible bulks. The RNAsimple Total RNA Extraction Kit (Tiangen, Beijing, China) was employed to extract total RNA from the leaf tissues according to the manufacturer’s instructions. To prepare the resistant and susceptible RNA pools, equal quantities of RNA from 30 homozygous resistant and 30 homozygous susceptible plants within the F2:3 families were combined. RNA quality was verified through agarose gel electrophoresis and spectrophotometric analysis. Subsequently, mRNA was isolated by poly(A)+ affinity purification using oligo(dT) 25 magnetic beads. Following 15 min–30 min binding at 18 °C–20 °C with gentle mixing, the beads were washed to deplete non-mRNA species. Eluted poly(A)+ RNA was fragmented either chemically (94 °C Mg2+ buffer) or enzymatically (RNase III, 37 °C) to generate 100 nt–300 nt fragments; then, reverse transcriptase and random hexamers were used to synthesize cDNA. The cDNA molecules were then processed through end-repair and adapter ligation reactions. The fragments that had been ligated were subjected to amplification through polymerase chain reaction (PCR) to construct the ultimate sequencing library, which was quantified and quality-controlled to meet sequencing requirements.

2.3. Genetic Analysis

For genetic analysis, F1 and F2 generations, as well as F2:3 families (randomly sampled 20 seeds per family) derived from the cross between L709 and LDN, along with the parental lines L709 and LDN, were sown and exposed to Bgt isolate E09 inoculation at the seedling stage. A total of 10 F1 plants, 246 F2 individuals, and their corresponding F2:3 families, as well as the parental lines L709 and LDN, were used for genetic analysis. ITs were scored on a 0–4 scale as described above [47]. Phenotypic data were subjected to goodness-of-fit analysis, and statistical analysis for phenotypic segregation ratios was conducted using SPSS Statistics version 16.0 (SPSS Inc., Chicago, IL, USA), with chi-square (χ2) tests applied to determine significant deviations between observed and expected genetic distributions.

2.4. RNA Extraction and Sequencing Library Setup for BSR-Seq

According to the identification results of powdery mildew resistance at the seedling stage of the F2:3 families of L709 and LDN, 30 homozygous resistant families and 30 homozygous susceptible families were selected to construct resistant and susceptible pools for transcriptomic sequencing analysis. The RNAsimple Total RNA Extraction Kit (Tiangen, Beijing, China) was employed to extract total RNA from young leaf tissues according to the manufacturer’s instructions. To prepare the resistant and susceptible RNA pools, equal quantities of RNA from 30 homozygous resistant and 30 homozygous susceptible plants within the F2:3 families were combined. RNA quality was verified through agarose gel electrophoresis and spectrophotometric analysis. Subsequently, mRNA was isolated by poly(A)+ affinity purification using oligo(dT) 25 magnetic beads. Following 15 min–30 min binding at 18–20 °C with gentle mixing, the beads were washed to deplete non-mRNA species. Eluted poly(A)+ RNA was fragmented either chemically (94 °C Mg2+ buffer) or enzymatically (RNase III, 37 °C) to generate 100 nt–300 nt fragments; then, reverse transcriptase and random hexamers were used to synthesize cDNA. The cDNA molecules were then processed through end-repair and adapter ligation reactions. The fragments that had been ligated were subjected to amplification through polymerase chain reaction (PCR) to construct the ultimate sequencing library, which was quantified and quality-controlled to meet sequencing requirements.

2.5. Sequencing and Bioinformatics Analysis

Sequencing was performed on the Illumina HiSeq 4000 platform at Tcuni Bioscience (Chengdu, China), a commercial genomics service provider. The process generated a large volume of short-read sequencing data. Raw data were subjected to quality control, including the removal of low-quality reads and trimming of adapter sequences. Related parameters include the following: (1) ILLUMINACLIP:TruSeq3-PE.Fa:2:30:10 for adapter sequence removal; (2) LEADING:20 and TRAILING:20 to trim low-quality bases (Q < 20) from read termini; (3) SLIDINGWINDOW:4:20 to scan reads with a 4-base window, trimming when average quality fell below 20; and (4) MINLEN:50 to discard reads shorter than 50 bp after processing. Filtered reads were mapped to the reference genome of T. turgidum cv. Svevo [48] using the STAR software. Read alignment was performed using STAR v2.3.0e with the following parameters: (1) genomeDir: /path/to/hg38 (Svevo v1.0 reference genome index); (2) runMode: alignReads; (3) outSAMtype: BAM SortedByCoordinate for coordinate-sorted alignments; (4) quantMode: TranscriptomeSAM to enable transcriptome quantification. SNPs were called using GATK v3.1-1 (depth ≥ 3, parent-pool consistent). Candidate regions were identified via 5 Mb sliding windows with 100 k permutations. Differentially expressed genes (DEGs) (FC ≥ 2, FDR < 0.01) were detected using EBSeq v3.5 software [49]. Functional enrichment were performed using clusterProfiler v4.6.0 in R package. Significant enrichment was observed for the plant–pathogen interaction pathway (ko04626; FDR < 0.05), among other defense-related pathways [49].

2.6. Candidate Interval Analysis and Gene Mapping

Genetic variant calling was performed using GATK [50], with SNPs filtered via Bcftools (v1.9) based on the following quality metrics: quality (QUAL) score > 30 and depth (DP) ≥ 5. To identify candidate genomic intervals, varBScore analysis [51] was applied to compare SNPs and insertion–deletion polymorphisms (Indels) between resistant and susceptible RNA pools. Additionally, Euclidean distance (ED) values of SNPs were evaluated using quantile-based thresholding, retaining the top 1% of SNPs to minimize noise from individual loci [52].
After identifying candidate intervals using ED analysis, Indels and SNPs within these intervals were searched in the resistant and susceptible bulks to design polymorphic markers. Using the 3.0 kb flanking sequences of each variant as templates, Indel primers were designed using the Primer Premier 5.0 software and the PrimerServer function on the WheatOmics platform (http://202.194.139.32/PrimerServer/) (accessed on 7 February 2025). All newly designed markers, along with previously reported markers in similar regions, were tested for polymorphisms between the resistant and susceptible parents and bulks. Subsequently, these markers were used to genotype the F2:3 families of L709 × LDN for gene mapping. After obtaining genotype data from marker analysis and phenotype data from disease resistance assessment of the segregating population, the linkage relationship between the markers and the Pm gene(s) in L709 was analyzed. A genetic linkage map was constructed by employing Mapmaker 3.0b [53], with a logarithm of odds score threshold set at 3.0.

2.7. Differential Expression Gene Analysis, Functional Annotation, and Enrichment Analysis of DEGs

Following BSR-Seq, the data analysis tool featureCounts v2.0.1 was employed to quantify gene or transcript expression levels by quantifying reads mapped to every gene. Statistical comparisons were carried out between the resistant and susceptible pools. EBSeq version 3.5 software was utilized to detect DEGs with a fold-change of at least 2 and a false discovery rate below 0.01. Following this, functional annotation of the identified DEGs was carried out using the Gene Ontology (GO, http://geneontology.org/) (accessed on 7 February 2025) and Kyoto Encyclopedia of Genes and Genomes (KEGG, https://www.kegg.jp/) (accessed on 7 February 2025) databases. An R package (clusterProfiler version 4.6.0, https://bioconductor.org/packages/release/bioc/html/clusterProfiler.html.) (accessed on 7 February 2025) developed for DEG analysis [54] was employed to support this annotation process. Through this step, the functional roles and biological pathways related to the DEGs could be determined. Lastly, an enrichment analysis was executed to investigate the biological processes, cellular structures, or molecular functions that occur in an over-represented manner within the set of DEGs.

2.8. Microscopic Observation of Powdery Mildew Resistance

The resistant parent L709 and susceptible parent LDN were grown under controlled conditions until reaching the one-leaf-one-heart developmental stage. Fresh spores of the Bgt isolate E09 were then inoculated onto the plants using the brush inoculation method. The environmental conditions at the time of inoculation were controlled at a temperature range of 18–22 °C and a humidity range of 60–80%. Leaf segments (2–3 cm in length) were harvested at eight time points: 0, 4, 12, 24, 36, 48, 72, and 120 h post inoculation (hpi). Collected tissues were immediately submerged in Carnoy’s fixative solution (anhydrous ethanol/glacial acetic acid = 3:1 v/v) for a minimum of 48 h to facilitate chlorophyll removal.
The leaves were then soaked in a 0.6% Coomassie Brilliant Blue solution (0.6 g Coomassie Brilliant Blue powder dissolved in 100 mL anhydrous ethanol) for 1 min, followed by thorough rinsing with water to remove excess dye (repeated rinsing was carried out until the dye was completely washed off). The leaves were placed face-up on a slide, and a drop of glycerol was added to prepare the slide samples. The growth morphology of powdery mildew spores was observed by using an Axioscope 5 microscope (ZEISS, Oberkochen, Germany). Images were acquired at a magnification of ×400.

2.9. RNA Extraction and qRT-PCR

First, we planted the disease-resistant parent L709 and the disease-susceptible parent LDN. When they reached the one-leaf stage, we collected the first leaves of L709 and LDN at eight time points: 0, 0.5, 2, 4, 12, 24, 48, and 72 hpi. Subsequently, RNA was extracted, and reverse transcription was performed to synthesize cDNA. Total RNA isolation was performed using the FastPure Universal Plant Total RNA Isolation Kit (Vazyme, Nanjing, China). Throughout the RNA extraction process, RNase-free equipment (Tiangen, Beijing, China) was required. In the process of reverse-transcribing RNA, the HiScript II® QRT SuperMix for qPCR (+gDNA wiper) kit was employed.
To characterize the expression profiles of resistance genes within the identified interval against Bgt isolates, qRT-PCR primers were designed using Primer Premier 5.0 software based on candidate gene sequences. qRT-PCR analysis was performed using the ChamQ Universal SYBR qPCR Master Mix (Vazyme, Nanjing, China), which contains SYBR Green I fluorescent dye and requires light protection during handling. The housekeeping gene TaActin was employed as an internal reference, and relative expression levels in L709 and LDN at multiple time points post E09 inoculation were calculated using the 2−ΔΔCT method [55]. Each reaction included three technical replicates.

2.10. Assessment of Linked Markers for MAS

To identify the availability of markers for MAS, tightly linked molecular markers (the molecular marker site is less than 5 cM away from the target gene) flanking the Pm locus in L709 were utilized to genotype L709 and 46 elite wheat cultivars collected from diverse agro-ecological zones in China. Markers capable of amplifying polymorphic bands between L709 and target wheat genotypes were validated for MAS in these genetic backgrounds. Otherwise, these markers could not be used to track the target Pm gene in those genetic backgrounds. In addition, 31 Pm gene donors were genotyped using these markers flanking the Pm locus in L709 to test whether the marker could distinguish between L709 and 31 donors.

3. Results

3.1. Evaluation and Genetic Analysis of Powdery Mildew Resistance in L709

Inoculation with Bgt isolate E09 revealed distinct phenotypic responses: cultivated emmer wheat accession L709 showed no visible symptoms and was immune with IT 0, whereas the susceptible parent LDN displayed a highly susceptible reaction (IT 4) (Figure 1A,B). All 10 F1 progenies derived from the cross L709 × LDN displayed resistant phenotypes with IT 0–1 (Table 1). A total of 246 F2 plants segregated with 183 resistant and 63 susceptible individuals, conforming to the expected 3:1 Mendelian monogenic ratio (χ2 = 0.05; p = 0.83). The results of the 246 F2:3 families further validate monogenic inheritance, yielding a phenotypic ratio of 60 homozygous resistant:123 segregating:63 homozygous susceptible families (χ2 = 0.07; p = 0.96) (Table 1). Collectively, these results indicate that resistance to Bgt isolate E09 in L709 is governed by a single dominant gene, provisionally designated PmL709.
Using resistance phenotyping data from Bgt isolate E09, ten homozygous resistant and ten homozygous susceptible F2:3 families were randomly selected from the L709 × LDN cross. These families were evaluated against eight additional Bgt isolates (E21, E23, E31, E32, F01, F02, F05, and F06) under controlled environmental conditions in eight independent growth chambers. The ten E09-resistant families consistently displayed broad-spectrum resistance against all tested Bgt isolates, whereas the ten E09-susceptible families remained uniformly susceptible (Table S1). These results indicate that the resistance of L709 to other Bgt isolates was also conferred by PmL709.

3.2. SNP Calling and Confirmation of Candidate Interval

BSR-Seq analysis generated high-quality sequence data for both resistant and susceptible bulks. The resistant bulk yielded 19.53 Gb of clean reads with a Q30 quality score exceeding 93% and a GC content ranging from 42 to 72% (Figure S1). Similarly, the susceptible bulk produced 21.67 Gb of clean data, featuring a Q30 score > 94% and a GC content of 41–70% (Figure S1). Following alignment, 130,844,322 sequence reads from the resistant bulk and 145,329,730 reads from the susceptible bulk were successfully mapped to the wild durum wheat (T. turgidum subsp. durum cv. Svevo) reference genome (svevo.v1), respectively (Figure S1). The high-quality sequence datasets were deemed suitable for downstream analyses. To reduce background noise, the original ED values were transformed by raising them to the 4th power and used as correlation values. The ED values were then fitted using a Loess curve. The selection of the threshold employs the quantile method, which involves sorting all fitted values in ascending order and selecting the ED values of SNP markers above 99% as the threshold during screening. Applying this threshold, only one estimated candidate region located on the terminal of chromosome arm 2BS (16.0–50.5 Mb) was detected based on the ED algorithm (Figure 2). This result indicates that PmL709 was located on chromosome arm 2BS.

3.3. Molecular Mapping of PmL709 and Prediction of Candidate Genes

To verify the mapping location, we specifically selected 12 reported SSR and Indel markers surrounding the region of interest on chromosome arm 2BS and developed ten Indel markers based on the results of the BSR-seq analysis. Twelve markers, including Xdw02, Xdw03, Xdw05, YTU95-04, YTU95-06, YTU95-08, Xdw10, Xdw11, YTU692B-094, YTU692B-102, XRE4, and WGGCI115 (Table S2), exhibited polymorphism between resistant and susceptible parents and bulks. Subsequently, these markers were used to genotype the 246 F2:3 families, resulting in the construction of a genetic linkage map for PmL709 (Figure 3A). Based on the linkage results, PmL709 was mapped within a 1.7 cM interval flanked by markers Xdw05/YTU95-04/YTU95-06/YTU95-08/Xdw10/Xdw11 and YTU692B-094 (Figure 3A). Based on the gene annotation results, a total of 100 genes with high confidence were identified within this interval. Among these genes, nine genes (including TRITD2Bv1G010130, TRITD2Bv1G010230, TRITD2Bv1G010660, TRITD2Bv1G010300, TRITD2Bv1G012060 TRITD2Bv1G010140, TRITD2Bv1G010240, TRITD2Bv1G011720, and TRITD2Bv1G012310) were associated with disease resistance and were considered as the candidate genes of PmL709.

3.4. Comparisons of PmL709 and Known Pm Genes on Chromosome Arm 2BS

Pm26, pm42, PmXQ-0508, Pm68, MlIW170, and PmWE99 genes have been reported on chromosome arm 2BS (Figure 3B). To identify the relationship between PmL709 and those genes, their donors with L709 were tested against 27 Bgt isolates to evaluate their resistance spectrum. The results show that L709 and TRI 1796 (Pm68) were resistant to all 27 isolates, while Pm26-40 (Pm26), B053299 (pm42), XQ-0508 (PmXQ-0508), IWM170 (MlIW170), and WE99 (PmWE99) were resistant to 25 of 27 (92.6%), 23 of 27 (85.2%), 17 of 27 (63.0%), 14 of 27 (51.9%), and 21 of 27 (77.8%) isolates, respectively. Therefore, L709 had a different resistance spectrum from the known Pm genes on 2BS, except for TRI 1796 (Pm68) (Table S3).

3.5. Discovery and Analysis of DEGs

Using BSR-Seq, a total of 3923 DEGs were identified between the two bulks, with 1846 down-regulated and 2077 up-regulated genes relative to the susceptible bulk (Figure 4A, Table S4). A GO analysis further revealed that these DEGs were mainly involved in three main domains: (i) molecular function, such as transmembrane transporter activity and chlorophyll binding, which are essential for the transport of molecules across membranes and the binding of chlorophyll, respectively; (ii) the cellular component, in particular the thylakoid, chloroplast thylakoid membrane, plastid thylakoid membrane, and integral components of the plasma membrane, which are crucial for photosynthesis and other cellular processes; and (iii) biological processes, e.g., photosynthesis (both light harvesting in photosystem I and light reactions), responses to various stimuli (such as acid chemicals, oxygen-containing compounds, water, and hormones), and responses to endogenous stimuli and water deprivation, suggesting that these genes could act as key mediators in activating defense responses against pathogenic infections (Figure 4B). To further elucidate metabolic pathways and characterize the molecular mechanisms underlying the biological processes that the DEGs may be involved in, a KEGG pathway enrichment analysis was conducted on the DEGs in the differential expression analysis (Figure 4C, Table S5). The results reveal several pathways associated with plant disease resistance. The phenylpropanoid and flavonoid biosynthesis pathways contribute to the production of antimicrobial compounds, while MAPK signaling and plant–pathogen interaction pathways play pivotal roles in defense mechanisms [56]. Furthermore, glutathione metabolism aids in alleviating oxidative stress during infection. These findings underscore the importance of metabolic and signaling pathways in bolstering plant immunity and stress tolerance.

3.6. Validation of Disease Resistance-Related Genes in Cultivated Emmer Wheat L709 via qRT-PCR

Upon further analysis, it was found that only 82 DEGs were situated within the candidate interval spanning from 21.82 to 25.94 Mb. These genes are hypothesized to be the essential candidates that play a role in the resistance against powdery mildew in accession L709. We investigated the expression profiles of nine DEGs located within this candidate interval at various time points (hpi) with Bgt isolate E09 in the resistant parent L709 and the susceptible parent LDN.
All of these genes were induced to be expressed by the Bgt invasion. The transcription levels of five genes, namely TRITD2Bv1G010130, TRITD2Bv1G010230, TRITD2Bv1G010660, TRITD2Bv1G010300, and TRITD2Bv1G012060, were higher in resistant parent L709 than susceptible parent LDN in most time points. Among the analyzed genes, TRITD2Bv1G010230 and TRITD2Bv1G010300 exhibited peak transcript levels at 2 hpi, while TRITD2Bv1G010130 and TRITD2Bv1G012060 peaked earlier at 0.5 hpi. In contrast, TRITD2Bv1G010660 showed a delayed peak expression at 48 hpi. Their expression levels ranged from 0 to 5-fold relative to the control (0 hpi). Notably, TRITD2Bv1G010140, TRITD2Bv1G010240, TRITD2Bv1G011720, and TRITD2Bv1G012310 displayed significantly lower expression in L709 compared to LDN (Figure 5, Table 2), suggesting potential negative regulatory mechanisms.

3.7. Molecular Markers for MAS

To facilitate the implementation of MAS targeting PmL709, seven flanking markers were evaluated for their utility in 46 susceptible wheat genotypes. Among them, marker YTU95-04 generated polymorphic amplicons between L709 and all 46 accessions, while the remaining six markers displayed polymorphism with most tested genotypes. These results indicate their suitability for MAS applications to track PmL709 during introgression into target genetic backgrounds (Figure 6, Table S6). These seven markers were also tested with 31 known Pm gene donors, including TRI 1796 (Pm68), XQ-0508 (PmXQ-0508), WE99 (PmWE99), and IWM170 (MlIW170), located on chromosome 2BS. The results show that some or all markers could effectively differentiate between these 31 donors and L709 (PmL709) (Table S7).

4. Discussion

The cultivated emmer wheat is a valuable gene donor for controlling wheat powdery mildew, which significantly reduces global wheat yields. As a progenitor of modern wheat, emmer wheat possesses abundant genetic diversity, which includes a large number of resistance genes/alleles that have been lost during the domestication and breeding of wheat cultivars [57,58]. Exploring the potential of emmer wheat for wheat breeding against powdery mildew could offer a robust solution to control this disease. In our lab, a cultivated emmer wheat accession, L709, demonstrated high resistance to powdery mildew in the field. To better use this resistance in breeding against powdery mildew, we dissected its genetic basis for this resistance and also assessed its breeding value in the present study.
Using genetic analysis, BSR-Seq, and molecular marker analysis, we accurately and rapidly identified a dominant Pm gene, PmL709, in cultivated emmer wheat L709 and localized it to a 1.7 cM genetic interval, corresponding to a 21.82–25.94 Mb physical interval on chromosome arm 2BS based on the reference genome of T. turgidum cv. Svevo. In previous studies, six Pm genes were reported on this chromosome arm from diversified gene donors, including Pm26 from TTD140 (T. turgidum var. dicoccoides) [39], pm42 from G-303-1M (T. turgidum var. dicoccoides) [40,41], PmXQ-0508 from XQ-0508 (T. aestivum L.) [40], Pm68 from TIR 1796 (T. turgidum L. var. durum Desf) [42], MlIW170 from IW170 (T. turgidum var. dicoccoides) [43], and PmWE99 from WE99 (Thinopyrum intermedium) [44]. Compared with those documented Pm genes, PmL709 (21.82–25.94 Mb) could be clearly distinguished from five of them based on their physical position: PmXQ-0508 (28.86–29.59 Mb), Pm26 (co-separated with Xcau516 at a locus similar to PmXQ-0508), pm42 (72.33–108.46 Mb), MlIW170 (30.10–30.59 Mb), and PmWE99 (unknown-108.46 Mb). Differential resistance spectrum results also provide evidence to support these differences between PmL709 and PmXQ-0508, Pm26, pm42, MlIW170, and PmWE99 (Table S3). However, the physical interval of PmL709 (21.82–25.94 Mb) overlapped that of Pm68 (24.44–26.64 Mb). We speculate that PmL709 might be a Pm68 allele or a new Pm gene closely linked with Pm68 due to the different species origin. To date, Pm68 has only been mapped to a physical interval of 2.20 Mb and has not been cloned. In the future, functional validation via allelism tests, gene cloning, and resistance mechanism dissection is required to establish their genetic relationships. Whether a Pm68 allele or a new Pm gene, our study provides valuable information to clarify this interval harboring valuable resistance genes, accelerating the cloning of this locus and breeding use against powdery mildew in production.
To further clarify the candidate interval and analyze the candidate genes/controlling genes, we also identified 3923 DEGs between resistant and susceptible bulks using BSR-Seq analysis, with 1846 being down-regulated and 2077 being up-regulated. We further classified these DEGs based on their functions through a GO analysis and highlighted the involvement of phenylpropanoid and flavonoid biosynthesis, MAPK signaling, and plant–pathogen interaction pathways using a KEGG pathway analysis, which are known to play critical roles in plant defense mechanisms [54]. All of these findings provide valuable information to understand the molecular mechanism of PmL709 in fighting Bgt invasion and to clone this gene. To further analyze the candidate of PmL709, we selected nine candidate genes associated with disease resistance within the PmL709 interval and profiled the resistance patterns after Bgt invasion. Five genes, including those encoding NBS-LRR proteins and receptor-like kinases, were up-regulated in L709 compared with the susceptible parent, suggesting their potential role in mediating the resistance to Bgt invasion [59]. This is consistent with previous research findings showing that the reported disease resistance genes in wheat and other species mainly encode two types of proteins: NLR proteins and kinase proteins. For example, genes including Pm1a, Pm2a, Pm3b/Pm8, Pm5e, Pm12, Pm21, Pm41, Pm60/MlIW172, and Pm69 are characterized as encoding NLR proteins [8,9,10,11], which serve as central mediators in plant defense responses. Pm24 encodes a tandem kinase protein, while Pm4 generates a chimeric polypeptide characterized by a pseudokinase domain (serine/threonine specificity), multiple C2 domains, and a transmembrane structural motif. Significantly, two cloned resistance genes, Pm38 encoding an ABC transporter and Pm46 encoding a hexose transporter, have been identified to confer broad-spectrum disease resistance against both powdery mildew and rust pathogens in wheat [13,14]. These results provide preliminary evidence for the functional roles of these candidate genes and highlight the need for further functional validation using techniques such as gene knockout or over-expression [60].
The ultimate objective of identifying disease-resistant genes is to integrate them into practical breeding applications. The implementation of cutting-edge molecular breeding techniques, including MAS, will allow resistance genes to be precisely introgressed into susceptible wheat cultivars. To enable the efficient introgression of PmL709 via MAS, we assessed the utility of seven molecular markers—comprising two novel markers and five previously reported markers—across 46 susceptible wheat cultivars in China. A polymorphism analysis revealed that all of these markers can discriminate L709 and the majority of tested genotypes, indicating their suitability for MAS-based detection of PmL709 during introgression into recipient wheat genotypes. In fact, we transferred PmL709 into several wheat cultivars and obtained the BC1F2 and F3 segregation populations. It is anticipated that PmL709 will realize its full potential through concurrent selection for disease resistance and agronomic traits in wheat breeding programs.

5. Conclusions

(1)
Cultivated emmer wheat accession L709 is resistant to 27 Bgt isolates and has extremely important breeding value.
(2)
The resistance to powdery mildew was conferred by the broad-spectrum resistance gene PmL709 in accession L709. PmL709 was delimited to a 1.7 cM genetic interval on chromosome arm 2BS, corresponding to a 21.82–25.94 Mb physical interval (svevo.v1).
(3)
DEGs, including TRITD2Bv1G010130, TRITD2Bv1G010230, TRITD2Bv1G010660, TRITD2Bv1G010300, TRITD2Bv1G012060, TRITD2Bv1G010140, TRITD2Bv1G010240, TRITD2Bv1G011720, and TRITD2Bv1G012310, located in the candidate interval, were induced to be expressed by Bgt invasion and could be candidates during the process of disease resistance response in L709.
(4)
The seven flanking markers of PmL709 (including Xdw05, YTU95-04, YTU95-06, YTU95-08, Xdw10, Xdw11, and YTU692B-094) can be used in MAS to track the PmL709 gene when transferring it to susceptible cultivars/lines.

Supplementary Materials

The following supporting information can be downloaded at https://www.mdpi.com/article/10.3390/agronomy15040980/s1, Table S1. Reaction patterns of 10 homozygous resistance and 10 homozygous susceptible F2:3 families to eight Blumeria graminis f. sp. tritici (Bgt) isolates. Table S2. Molecular markers and their primer sequences used in this study. Table S3. Comparative responses of L709 and wheat genotypes with known powdery mildew resistance genes on chromosome arm 2BS to 27 Blumeria graminis f. sp. tritici (Bgt) isolates with different virulence. Table S4. Differentially expressed genes (DEGs) between resistant and susceptible bulks of L709 × Langdon (LDN). Table S5. Information about the detailed Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway for differentially expressed genes (DEGs) on 14 emmer wheat chromosomes (2n = 14, AABB). Table S6. Validation of PmL709-linked markers on 46 Chinese wheat cultivars/breeding lines in marker-assisted selection (MAS) breeding. Table S7. Validation of PmL709-linked markers in 31 known powdery mildew resistance gene donors. Figure S1. Sequencing quality analysis of bulked segregant RNA sequencing (BSR-Seq) on resistance and susceptible bulks of L709 × Langdon (LDN).

Author Contributions

Conceptualization, Y.J. and P.M.; Data Curation, R.L., N.Y. and X.L.; Formal Analysis, J.Z., J.L. and Y.L.; Funding Acquisition, Y.J., N.Y., J.W. and P.M.; Investigation, J.Z., J.L. and Y.L.; Methodology, H.X., X.S. and P.M.; Resources, J.W.; Supervision, H.X., X.S. and X.L.; Validation, Y.J., N.Y., H.X., X.S., J.W., X.L., J.Z., J.L. and Y.L.; Visualization, R.L. and N.Y.; Writing—Original Draft, R.L., Y.J. and X.L.; Writing—Review and Editing, P.M. All authors have read and agreed to the published version of the manuscript.

Funding

This research was financially supported by the Taishan Scholar Foundation of Shandong Province (TSCY202211144), the National Natural Science Foundation of China (32301923), Shandong Province Key R&D Plan (2024LZGCQY012), and the Natural Science Foundation of Shandong Province (ZR2023QC203, ZR2023QC292, and ZR2024MC039).

Institutional Review Board Statement

Not applicable.

Data Availability Statement

The authors confirm that the data supporting the findings of this study are available within the article or its Supplementary Materials.

Conflicts of Interest

The author Wang Jiangchun was employed by Shandong Zhongnong Tiantai Seed Industry Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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Figure 1. Phenotype analysis of L709 and susceptible parent Langdon (LDN) after inoculation with Blumeria graminis f. sp. tritici (Bgt) isolate E09. (A) Phenotype of L709, LDN, and part of F2:3 plants about 21 days after inoculation with Bgt isolate E09; (B) infection process of Bgt isolate E09 on leaves of L709 and LDN. Wheat leaf samples were taken at different hours post inoculation (hpi) for coomassie blue staining. Scale bar: 200 μm.
Figure 1. Phenotype analysis of L709 and susceptible parent Langdon (LDN) after inoculation with Blumeria graminis f. sp. tritici (Bgt) isolate E09. (A) Phenotype of L709, LDN, and part of F2:3 plants about 21 days after inoculation with Bgt isolate E09; (B) infection process of Bgt isolate E09 on leaves of L709 and LDN. Wheat leaf samples were taken at different hours post inoculation (hpi) for coomassie blue staining. Scale bar: 200 μm.
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Figure 2. The identification of the candidate region for PmL709 using the Euclidean distance (ED) algorithm. 1A–7B indicate wheat chromosomes; fitted: the quantile method was used to select the threshold of the ED4 fitting loess curve; unfitted: unfitted ED4.
Figure 2. The identification of the candidate region for PmL709 using the Euclidean distance (ED) algorithm. 1A–7B indicate wheat chromosomes; fitted: the quantile method was used to select the threshold of the ED4 fitting loess curve; unfitted: unfitted ED4.
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Figure 3. The mapping of Pm709 and its positional relationship with the reported Pm genes on the same chromosome arm. (A) A linkage map of PmL709 using F2:3 families from cross L709 × Langdon. (B) The physical positions of reported Pm genes on wheat chromosome 2BS (based on the flanking markers of each gene referring to the genome of IWGSC RefSeq v2.1). Note: The left side is the genetic distance in cM; the right side is the physical distance in bp; the red genes represent genes located on chromosome 2BS [39,40,41].
Figure 3. The mapping of Pm709 and its positional relationship with the reported Pm genes on the same chromosome arm. (A) A linkage map of PmL709 using F2:3 families from cross L709 × Langdon. (B) The physical positions of reported Pm genes on wheat chromosome 2BS (based on the flanking markers of each gene referring to the genome of IWGSC RefSeq v2.1). Note: The left side is the genetic distance in cM; the right side is the physical distance in bp; the red genes represent genes located on chromosome 2BS [39,40,41].
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Figure 4. The distribution and functional classification of differentially expressed genes (DEGs) based on the bulked segregant RNA sequencing (BSR-Seq) of the resistance and susceptible bulks of L709 × Langdon. (A) The distribution of the DEGs using a volcano plot. (B). A Gene Ontology (GO) analysis of the DEGs. (C) A Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the DEGs.
Figure 4. The distribution and functional classification of differentially expressed genes (DEGs) based on the bulked segregant RNA sequencing (BSR-Seq) of the resistance and susceptible bulks of L709 × Langdon. (A) The distribution of the DEGs using a volcano plot. (B). A Gene Ontology (GO) analysis of the DEGs. (C) A Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the DEGs.
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Figure 5. qRT-PCR analysis of potential candidate genes of PmL709 (TRITD2Bv1G010130, TRITD2Bv1G010140, TRITD2Bv1G010230, TRITD2Bv1G010240, TRITD2Bv1G010300, TRITD2Bv1G010660, TRITD2Bv1G011720, TRITD2Bv1G012060, and TRITD2Bv1G012310) in resistant parent L709 and susceptible parent Langdon after inoculating with Blumeria graminis f. sp. tritici (Bgt) isolate E09 at 0, 0.5, 2, 4, 12, 24, 48, and 72 h post inoculation (hpi). Asterisks indicate significant differences (t tests) between L709 and LDN at each time point (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns: not significant).
Figure 5. qRT-PCR analysis of potential candidate genes of PmL709 (TRITD2Bv1G010130, TRITD2Bv1G010140, TRITD2Bv1G010230, TRITD2Bv1G010240, TRITD2Bv1G010300, TRITD2Bv1G010660, TRITD2Bv1G011720, TRITD2Bv1G012060, and TRITD2Bv1G012310) in resistant parent L709 and susceptible parent Langdon after inoculating with Blumeria graminis f. sp. tritici (Bgt) isolate E09 at 0, 0.5, 2, 4, 12, 24, 48, and 72 h post inoculation (hpi). Asterisks indicate significant differences (t tests) between L709 and LDN at each time point (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns: not significant).
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Figure 6. Amplification results of molecular markers YTU692B-094 and YTU95-04 in 15 representative wheat cultivars/breeding lines. M, pUC19/MspI; lane 1, resistant parent L709; lane 2, susceptible parent Langdon. Lanes 3–17 indicate partial powdery mildew-susceptible wheat cultivars/breeding lines derived from multiple major wheat-producing areas and possessing excellent agronomic and yield traits: Yannong 5158, Saidemai 16, Shengmai 116, Yunchengxiaomai, Tai 4123, Nongda 4538, Zhengmai 16, Xinmai 23, Yumai 49, Shimai 15, Shinong 086, Taimai 101, Huaimai 23, Chang 6878, and Jimai 19. Red arrow indicates polymorphic band.
Figure 6. Amplification results of molecular markers YTU692B-094 and YTU95-04 in 15 representative wheat cultivars/breeding lines. M, pUC19/MspI; lane 1, resistant parent L709; lane 2, susceptible parent Langdon. Lanes 3–17 indicate partial powdery mildew-susceptible wheat cultivars/breeding lines derived from multiple major wheat-producing areas and possessing excellent agronomic and yield traits: Yannong 5158, Saidemai 16, Shengmai 116, Yunchengxiaomai, Tai 4123, Nongda 4538, Zhengmai 16, Xinmai 23, Yumai 49, Shimai 15, Shinong 086, Taimai 101, Huaimai 23, Chang 6878, and Jimai 19. Red arrow indicates polymorphic band.
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Table 1. Segregation ratios of F2 and F2:3 generations of cross L709 × Langdon (LDN) following inoculation with Blumeria graminis f. sp. tritici (Bgt) isolate E09 at seedling stage.
Table 1. Segregation ratios of F2 and F2:3 generations of cross L709 × Langdon (LDN) following inoculation with Blumeria graminis f. sp. tritici (Bgt) isolate E09 at seedling stage.
Parents and CrossGenerationObserved RatioExpected Ratioχ2p
L709RPR:S = 10:0
LDNSPR:S = 0:10
L709 × LDN F1F1R:S = 10: 0
L709 × LDN F2F2R:S = 183:633:10.050.83
L709 × LDN F2:3F2:3HR:Seg:HS = 60:123:631:2:10.070.96
Note: RP, resistant parent; SP, susceptible parent. R, resistant; S, susceptible; HR, homozygous resistant; Seg, segregating; HS, homozygous susceptible.
Table 2. Gene annotation of disease resistance-related genes in candidate interval of wheat powdery mildew resistance gene PmL709.
Table 2. Gene annotation of disease resistance-related genes in candidate interval of wheat powdery mildew resistance gene PmL709.
No.GenesPhysical Genomic LocationFunctional Annotation
1TRITD2Bv1G010130chr2B:21983992..21987186Disease resistance protein
2TRITD2Bv1G010140chr2B:21994145..21997744Disease resistance protein (NBS-LRR class) family
3TRITD2Bv1G010230chr2B:22398122..22400896NBS-LRR-like resistance protein
4TRITD2Bv1G010240chr2B:22456742..22460374NBS-LRR-like resistance protein
5TRITD2Bv1G010300chr2B:22629909..22631735Disease resistance protein (TIR-NBS-LRR class)
6TRITD2Bv1G010660chr2B:23210823..23214726Receptor protein kinase
7TRITD2Bv1G011720chr2B:24749117..24754598Disease resistance protein (TIR-NBS-LRR class)
8TRITD2Bv1G012060chr2B:25098753..25102541Receptor-like protein kinase
9TRITD2Bv1G012310chr2B:25474820..25483319NBS-LRR class disease resistance protein
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MDPI and ACS Style

Liu, R.; Jin, Y.; Yu, N.; Xu, H.; Sun, X.; Wang, J.; Liu, X.; Zhang, J.; Li, J.; Li, Y.; et al. Genetic Dissection of the Powdery Mildew Resistance in a Cultivated Emmer Wheat Accession. Agronomy 2025, 15, 980. https://doi.org/10.3390/agronomy15040980

AMA Style

Liu R, Jin Y, Yu N, Xu H, Sun X, Wang J, Liu X, Zhang J, Li J, Li Y, et al. Genetic Dissection of the Powdery Mildew Resistance in a Cultivated Emmer Wheat Accession. Agronomy. 2025; 15(4):980. https://doi.org/10.3390/agronomy15040980

Chicago/Turabian Style

Liu, Ruishan, Yuli Jin, Ningning Yu, Hongxing Xu, Xusheng Sun, Jiangchun Wang, Xueqing Liu, Jiadong Zhang, Jiatong Li, Yaoxue Li, and et al. 2025. "Genetic Dissection of the Powdery Mildew Resistance in a Cultivated Emmer Wheat Accession" Agronomy 15, no. 4: 980. https://doi.org/10.3390/agronomy15040980

APA Style

Liu, R., Jin, Y., Yu, N., Xu, H., Sun, X., Wang, J., Liu, X., Zhang, J., Li, J., Li, Y., & Ma, P. (2025). Genetic Dissection of the Powdery Mildew Resistance in a Cultivated Emmer Wheat Accession. Agronomy, 15(4), 980. https://doi.org/10.3390/agronomy15040980

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