Molecular markers tightly linked to the wheat powdery mildew resistance gene PmDR691 and methods of use thereof

Abstract

The application belongs to the field of agricultural biology genes and relates to a wheat powdery mildew resistance gene PmDR691 The application relates to a closely linked molecular marker, application and method thereof. The molecular marker is located on a 7BL chromosome of wheat and is at 696934274bp. A primer set for identifying the molecular marker comprises an upstream primer HENU1060-F with a nucleotide sequence as shown in SEQ ID No. 1; and a downstream primer HENU1060-R with a nucleotide sequence as shown in SEQ ID No. 2. The primer provided by the application can be used to detect whether a wheat breeding material carries a powdery mildew resistance gene before powdery mildew does not occur in a wheat seedling stage PmDR691 The marker can be used to detect and select a variety carrying the powdery mildew resistance gene PmDR691 in a targeted manner. The marker is not only fast, accurate and cost-saving, but also improves breeding efficiency and speeds up the breeding process.

Description

Technical Field

[0001] This invention belongs to the field of agricultural biogenetics and relates to wheat genetics and breeding. Background Technology

[0002] Wheat is a widely cultivated and important food crop worldwide, playing a crucial role in agricultural production and food security in various countries. However, it is susceptible to damage from powdery mildew (Fungi brevicornu). Blumeria graminis f. sp. tritici Powdery mildew, caused by wheat, is prevalent in wheat-growing areas worldwide. In years of outbreaks, it can lead to yield losses of more than 50%, seriously threatening food security (Appels et al. Shifting the limits in wheat research and breeding using afully annotated reference genome. Science, 2018, 361:7191).

[0003] Due to the frequent genetic variation and rapid evolution of wheat powdery mildew pathogens, many widely used resistance genes often fail to maintain effective resistance in long-term production. Therefore, continuously discovering new powdery mildew resistance genes and rationally utilizing them for variety improvement through molecular marker-assisted selection and other methods has become the most economical, sustainable, and environmentally friendly strategy for controlling this disease. Breeding and promoting resistant varieties can not only reduce the use of chemical pesticides but also fundamentally improve the stability and safety of wheat production.

[0004] To date, over 100 genes resistant to powdery mildew have been identified, involving more than 80 resistance loci. Of these, over 90 genes have been formally named. Pm1-Pm71 It is worth noting that among these genes, besides... Pm5, Pm9, Pm26, Pm42, Pm47, Pm50, Pm58 and Pm61 Apart from recessive genes, most other powdery mildew resistance genes are controlled by dominant genes (Yahiaoui et al. Independent evolution of functional...). Pm3 resistancegenes in wild tetraploid wheat and domesticated bread wheat. The PlantJournal, 2009, 57: 846-856). Pm21This is currently the most resistant and broadest-spectrum powdery mildew resistance gene, widely used in wheat-growing areas of eastern my country. However, powdery mildew fungi are highly dynamic, capable of co-evolving with wheat, and their strains mutate very rapidly. Controlling a single resistance gene can easily lead to loss of resistance and a narrow genetic base for wheat varieties. Therefore, continuously discovering new powdery mildew resistance gene resources and broadening the genetic base of resistance has become a key technical challenge in current wheat disease resistance breeding.

[0005] Based on the origin of wheat powdery mildew resistance genes, they can be broadly classified into the following three categories. The primary wheat gene bank covers different lines and varieties of hexaploid common wheat (AABBDD), including local varieties and ancestral species. Within the primary wheat gene bank, a total of 48 officially named powdery mildew resistance genes and their alleles have been identified, such as... Pm1, Pm3, Pm24, Pm48, Pm65 Etc. Secondary genetic resources of wheat encompass species with one or two genomes closely related to common wheat. According to current official records, these species contain at least 35 known wheat powdery mildew resistance genes: for example... Pm3h, Pm68 (He et al. Characterization of Pm68 A new powdery mildew resistance gene on chromosome 2BS of Greek durumwheat TRI1796. Theoretical and Applied Genetics, 2021, 134:53-62) is derived from tetraploid durum wheat ( Triticum durum (AABB) Pm60 Derived from diploid Urartu wheat ( Triticum urartu (AA) Pm12, Pm32, Pm53 Source: diploid *Aegilops spp.* (Aegilops spp.) Aegilops speltoides These genes originate from various closely related wheat species, providing a rich selection space for breeding wheat resistant to powdery mildew. Using advanced techniques such as distant hybridization and chromosome engineering, these superior genes can be effectively introduced into common wheat to improve its resistance to powdery mildew (Wang et al. Fighting wheat powdery mildew: from genes to fields. Theoretical and Applied Genetics, 2023, 136: 196). The wheat tertiary gene pool consists of other closely related wheat species outside the primary and secondary gene pools, with the genus *Rhizophora* being frequently utilized. Secale ), genus Triticum ( Daspyrum ) and the genus *Leymus* ( Thinopyrum (Sun et al.) Pm61A recessive gene for resistance to topowdery mildew in wheat landrace (Xuxusanyuehuang identified by comparative genomics analysis. Theoretical and Applied Genetics, 2018, 131: 2085-2097). Among numerous closely related species, durum wheat, as an important cultivated species closely related to common wheat, possesses unique research and utilization value for its powdery mildew resistance gene resources.

[0006] durum wheat ( Triticum durum It is an important tetraploid cultivated wheat, possessing an AABB genome, and is phylogenetically related to common wheat ( Triticum aestivum The two wheat species are closely related. Therefore, in the process of genetic improvement, superior genes derived from durum wheat are more easily introduced into the common wheat background through conventional hybridization and backcrossing, exhibiting higher genetic compatibility and breeding efficiency compared to more distantly related species. Previous studies have shown that some powdery mildew resistance genes have been identified from durum wheat materials and successfully applied to the disease resistance improvement of common wheat, demonstrating their practical value in the discovery of resistance genes, such as the recently published gene from the durum wheat line TRI1796. Pm68 Two independent introgression lines have been developed by introducing the superior Chinese wheat variety Yangmai 158 through hybridization, and both lines have shown positive effects on seedling and mature plant growth. BgtYZ01 Expression of resistance (He et al. An NLR pair in the Pm68 (Locus confers powdery mildew resistance indurum and common wheat. Nature Communications, 2025, 16: 9039). Durum wheat germplasm resources are abundant and genetically diverse, containing a large number of powdery mildew resistance genes that have not yet been systematically explored and utilized. With the development of molecular marker technology and gene mapping technology, in-depth exploration and precise localization of durum wheat disease resistance resources will not only help broaden the genetic basis of wheat disease resistance but also provide an important gene source for breeding new varieties with durable resistance and a broad resistance spectrum. Therefore, durum wheat can be regarded as an important resource pool of wheat powdery mildew resistance genes and a potential source of superior resistance. Summary of the Invention

[0007] This invention proposes a gene for wheat resistance to powdery mildew. PmDR691 Tightly linked molecular markers and their applications and methods provide guidance for breeding new wheat varieties resistant to powdery mildew.

[0008] The technical solution of this invention is implemented as follows: This invention provides a wheat powdery mildew resistance gene. PmDR691 Molecular markers HENU1060 This molecular marker is related to genes PmDR691 The genetic distance is only 4 cM, located on the wheat 7BL chromosome at position 696934274bp.

[0009] It also provides a gene for amplifying powdery mildew resistance in durum wheat. PmDR691 Primers for tightly linked molecular markers, wherein the molecular markers are codominant SSR markers. HENU1060 Its primers include upstream primers. HENU1060-F and downstream primers HENU1060-R .

[0010] The molecular marker HENU1060 The upstream primer is HENU1060-F Its nucleotide sequence is: 5'-TGATGTCGTGTCATTGTGGAAAT-3', as shown in SEQ ID No. 1; The molecular marker HENU1060 The downstream primer is HENU1060-R Its nucleotide sequence is: 5'-ATCATGAGACGGACGCTTGG-3', as shown in SEQ ID No. 2.

[0011] A kit containing the above primers is also provided.

[0012] The amplified fruncal wheat powdery mildew resistance gene provided by this invention PmDR691 Primers for tightly linked molecular markers in genes PmDR691 The detection, identification, and auxiliary identification of wheat powdery mildew resistance traits, as well as its application in molecular marker-assisted breeding.

[0013] wheat powdery mildew resistance gene PmDR691 It is located on wheat chromosome 7BL (692,840,898-697,934,274bp).

[0014] This invention also provides a method for detecting whether a wheat sample carries a flint wheat powdery mildew resistance gene. PmDR691 The method includes the following steps: (1) Extract genomic DNA from the wheat sample to be tested; (2) PCR amplification of genomic DNA from wheat samples to be tested was performed using molecular marker primers to obtain amplification products; the molecular marker primers included upstream primers. HENU1060-F and downstream primers HENU1060-R ; The molecular marker HENU1060 The upstream primer is HENU1060-F Its nucleotide sequence is: 5'-TGATGTCGTGTCATTGTGGAAAT-3', as shown in SEQ ID No. 1; The downstream primer of the molecular marker HENU1060 is HENU1060-R Its nucleotide sequence is: 5'-ATCATGAGACGGACGCTTGG-3', as shown in SEQ ID No. 2; (3) Electrophoresis and detection of the amplified products. If a 616 bp tightly linked specific band can be amplified, it indicates that the wheat sample to be tested carries the flint wheat powdery mildew resistance gene. PmDR691 Otherwise, the wheat sample to be tested does not carry the powdery mildew resistance gene in durum wheat. PmDR691 .

[0015] The PCR amplification system in step (2) of the method is 10 μL, including: 1.0 μL of 50 ng / μL wheat genomic DNA, 4 μL of PCR Master Mix, 0.5 μL of 5 μM upstream primer, 0.5 μL of 5 μM downstream primer, and 4 μL of sterile deionized water.

[0016] The PCR amplification procedure in step (2) of the method is as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 15 s, 60℃ annealing for 15 s, extension for 40 s, 34 cycles; 72℃ extension for 10 min; storage at 12℃.

[0017] The electrophoresis procedure for the amplification product in step (3) of the method is as follows: electrophoresis is performed on a non-denaturing polyacrylamide gel with a mass-volume percentage concentration of 8%. The amplification product is mixed with 2 μL of 6× loading buffer, and 1.0 μL of the mixture is loaded. Electrophoresis is performed at a constant voltage of 180 V for 1.5-2 h. After staining with silver nitrate, the sample is photographed.

[0018] The present invention has the following beneficial effects: 1. The durum wheat variety Jing DR691 used in this study is immune to powdery mildew at both the seedling and mature stages. Genetic analysis and molecular marker detection of powdery mildew resistance at the seedling stage show that the resistance of Jing DR691 durum wheat seedlings to different types of wheat powdery mildew is caused by a pair of recessive genes. PmDR691 This novel wheat powdery mildew resistance gene / allele, located on wheat chromosome 7BL, is controlled by this gene. The wheat powdery mildew resistance gene provided by this invention... PmDR691 Molecular markers HENU1060 After genetic segregation population testing, and with genes PmDR691The genetic distance is only 4 cM, which is similar to the gene. PmDR691 Tight linkage allows for accurate detection of genes. PmDR691 Genetic mapping of large populations, applied to genes PmDR691 Fine-grained localization and map-based cloning.

[0019] 2. This invention provides a gene for resistance to wheat powdery mildew. PmDR691 Primers for tightly linked molecular markers and their applications not only enable rapid and accurate screening of target varieties, unaffected by the environment, and with clear selection targets, but also save production costs and greatly improve the efficiency and quality of selecting high-quality powdery mildew-resistant wheat varieties or lines. Attached Figure Description

[0020] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0021] Figure 1 molecular markers HENU1060 Results of segregating populations of durum wheat DR691 and Langdon and their hybrid progeny. In the figure: M: D2000; 1: Durum wheat DR691 (powdery mildew resistant variety, carrying the powdery mildew resistance gene). PmDR691 ); 2: Langdon durum wheat (susceptible to powdery mildew); 3: 20 families mixed with resistance; 4: 20 families mixed with susceptibility; 5-24: F1 generation formed by crossing durum wheat Jing DR691 and durum wheat Langdon. 2:3 Family lineage, of which 5-9: homozygous disease-resistant F1 cells 2:3 Family history, 10-19: F strains of immune system separation 2:3 Family history, 20-24: Homozygous F 2:3 Family lineage; white arrows indicate areas where genes can be traced. PmDR691 Specific bands.

[0022] Figure 2 The response of the parents DR691, Langdon and hybrid F1 progeny to strain E09. Detailed Implementation

[0023] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0024] Unless otherwise specified, the experimental methods used in the following experimental examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0025] Example 1: Wheat powdery mildew resistance gene PmDR691 Molecular markers HENU1060 Primer development 1. Materials Durum wheat varieties Jing DR691 and Langdon exhibit resistance and susceptibility to wheat powdery mildew, respectively. Crossing Jing DR691 and Langdon resulted in the F1 generation being self-crossed to obtain the F2 population and corresponding F1 generation. 2:3 The family tree is shown in Tables 1 and 2.

[0026] Table 1 shows the powdery mildew resistance phenotypes of plant materials.

[0027]

[0028] Table 2. DR691 and Langdon, and their F2 and F3 2:3 Segregation ratio of Bgt physiological race E09 inoculated at the seedling stage

[0029]

[0030] 2. Extraction of wheat genomic DNA The CTAB method for extracting wheat genomic DNA is as follows: (1) Cut fresh wheat leaves and grind them with liquid nitrogen. Take about 0.5 g and put it into a 2.0 mL centrifuge tube.

[0031] (2) Add 600 μL of CTAB extraction solution, incubate in a water bath at 65℃ for 40 min, and mix thoroughly every 10 min during the process.

[0032] (3) Add 600 μL of chloroform-isoamyl alcohol (24:1, v / v) and shake gently on a shaker for 15 min.

[0033] (4) Centrifuge at 12,000 rpm for 15 min, take the supernatant into another 2 mL centrifuge tube, add 3 times the volume of pre-cooled anhydrous ethanol, and place in a -20℃ refrigerator for precipitation for 60 min.

[0034] (5) Pick out the flocculent DNA precipitate and wash twice with 75% ethanol after pre-cooling.

[0035] (6) Pick out the DNA precipitate and place it in a 1.5 mL centrifuge tube. Let it air dry in the room.

[0036] (7) Add 60 μL of TE buffer (100 mM Tris-HCl, 10 mM EDTA). pH = 8.0) Dissolve the DNA precipitate to prepare a DNA storage solution.

[0037] (8) Dilute the DNA storage solution with ultrapure water to 20-50 ng / μL as a working solution for later use.

[0038] 3. Identification of powdery mildew resistance in wheat seedlings and genetic analysis of disease resistance The disease-resistant parental durum wheat Jing DR691, the disease-susceptible parental durum wheat Langdon, F1 hybrids, F2 populations, and F1 hybrids were compared. 2:3 The families were planted in 128-cell trays (2×2cm), with 5 seeds sown in each cell. Both the parents and F1 strains had 20 seeds each. 2:3 Twenty seeds were sown from each family, while the susceptible control Zhoumai 18 was randomly sown and tagged for identification. After sowing, growth conditions were controlled as a 14-hour light / 10-hour dark cycle, temperature 20℃, and relative humidity 50-80%. When seedlings reached the one-leaf stage, they were inoculated with wheat powdery mildew strain E09. For the first 24 hours after inoculation, conditions were controlled as darkness, temperature 20℃, and relative humidity 100%. Afterward, conditions were controlled as a 14-hour light / 10-hour dark cycle, temperature 18-22℃, and relative humidity 100%. When the first leaf of the susceptible control Zhoumai 18 showed full disease, phenotypes were recorded according to a 0-4 grade standard, where 0-2 was considered resistance and 3-4 was susceptibility.

[0039] The investigation results showed that durum wheat variety Jing DR691 was immune to strain E09, while durum wheat Langdon (LDN) was highly susceptible to strain E09. Crossing Jing DR691 with Langdon resulted in the F1 generation being susceptible to strain E09. Seedling resistance assessment of 379 F2 plants revealed 83 resistant plants and 296 susceptible plants, conforming to a 1:3 resistance-susceptibility segregation ratio. Seedling resistance assessment of 219 families derived from the F2 generation showed 49 homozygous resistant families, 116 heterozygous families, and 54 homozygous susceptible families, conforming to a 1:2:1 resistance-susceptibility segregation ratio. Based on the seedling resistance genetic analysis, the resistance of durum wheat Jing DR691 to strain E09 is controlled by a single recessive gene, which is tentatively named... PmDR691 .

[0040] 4. Preliminary molecular marker localization of the powdery mildew resistance gene in durum wheat Jing DR691 Based on phenotypic identification results, 20 homozygous resistant families and 20 homozygous susceptible families were selected to construct resistant and susceptible pools, respectively. Polymorphism detection was performed on durum wheat DR691 and Langdon, as well as the resistant and susceptible pools, using genome-wide uniformly distributed molecular markers. Ten pairs of markers showed consistent polymorphism in the resistant and susceptible parents and the resistant-susceptible pools. Subsequently, these markers were used to analyze the F1 generation of 219 durum wheat DR691 × Langdon durum wheat lines. 2:3 Genotyping of families, gene PmDR691 Preliminary localization to wheat chromosome 7BL.

[0041] 5. Genes PmDR691 Development of tightly linked molecular markers Based on the sequence information of the tetraploid wheat reference genome, *Durum wheat cv. Svevo* genome, Simple Sequence Repeat (SSR) markers were designed using Primer 5.0 software to target the F1 sequence of durum wheat *Jing DR691* × durum wheat *Langdon*. 2:3 Genotyping of the family pedigree yielded results related to the genes. PmDR691 Closely linked SSR tags HENU1060 The genetic distance is only 4 cM.

[0042] Molecular markers HENU1060 The primers consist of one upstream primer and one downstream primer: upstream primer HENU1060-F nucleotide sequence: 5'-TGATGTCGTGTCATTGTGGAAAT-3' (as shown in SEQ ID No. 1); Downstream primer HENU1060-R nucleotide sequence: 5'-ATCATGAGACGGACGCTTGG-3' (as shown in SEQ ID No. 2).

[0043] The PCR amplification system was 10 μL, comprising: 1.0 μL of 50 ng / μL wheat genomic DNA, 4 μL of PCR Master Mix, 0.5 μL of 5 μM upstream primer, 0.5 μL of 5 μM downstream primer, and 4 μL of sterile deionized water.

[0044] The PCR amplification program was as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 15 s, 60℃ annealing for 15 s, extension for 40 s, 34 cycles; 72℃ extension for 10 min; storage at 12℃.

[0045] The electrophoretic separation procedure for the amplified products is as follows: electrophoresis is performed on a non-denaturing polyacrylamide gel with a mass-volume percentage concentration of 8%. The amplified products are mixed with 2 μL of 6× loading buffer, and 1.0 μL of the mixture is loaded. Electrophoresis is performed at a constant voltage of 180V for 1.5-2 h. The gel is then stained with silver nitrate and photographed.

[0046] A specific 616 bp tightly linked band was amplified, indicating that the wheat sample carries the powdery mildew resistance gene. PmDR691 No specific band of 616 bp tightly linked was amplified, indicating that the wheat sample did not carry the gene for wheat resistance to powdery mildew. PmDR691 .

[0047] Example 2: Wheat powdery mildew resistance gene PmDR691 Molecular markers HENU1060 Application of primers Using wheat powdery mildew resistance genes PmDR691 Molecular markers HENU1060 Primer amplification detection of durum wheat DR691, durum wheat Langdon, and F1 hybrids formed by their crosses 2:3 The DNA extraction method for materials from the family lineage was the same as in Example 1.

[0048] Samples to be tested: durum wheat DR691, durum wheat Langdon, and their hybrids F1 2:3 Family lineage.

[0049] Genomic DNA was extracted from the above materials and used as a PCR amplification template, utilizing the molecular markers developed in this invention. HENU1060 Amplification was performed using primers: Nucleotide sequence of upstream primer HENU1060-F: 5'-TGATGTCGTGTCATTGTGGAAAT-3' (as shown in SEQ ID No. 1); The nucleotide sequence of the downstream primer HENU1060-R is as follows: 5'-ATCATGAGACGGACGCTTGG-3' (as shown in SEQ ID No. 2).

[0050] The PCR amplification system was 10 μL, comprising: 1.0 μL of 50 ng / μL wheat genomic DNA, 4 μL of PCR Master Mix, 0.5 μL of 5 μM upstream primer, 0.5 μL of 5 μM downstream primer, and 4 μL of sterile deionized water.

[0051] The PCR amplification procedure is as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 15 s, 55℃ annealing for 15 s, extension for 40 s, 34 cycles; 72℃ extension for 10 min; storage at 12℃.

[0052] The electrophoretic separation procedure for the amplified products is as follows: electrophoresis is performed on a non-denaturing polyacrylamide gel with a mass-volume percentage concentration of 8%. The amplified products are mixed with 2 μL of 6× loading buffer, and 1.0 μL of the mixture is loaded. Electrophoresis is performed at a constant voltage of 180V for 1.5-2 h. The mixture is then stained with silver nitrate and photographed.

[0053] Molecular marker detection results are shown below Figure 1 . Figure 1 For marking HENU1060 F derived from durum wheat DR691 × durum wheat Langdon 2:3 Partial amplification results from the family. In the figure: M: D2000; 1: Durum wheat Jing DR691 (a powdery mildew-resistant variety carrying the powdery mildew resistance gene). PmDR691 ); 2: Langdon durum wheat (susceptible to powdery mildew); 3: 20 families mixed with resistance; 4: 20 families mixed with susceptibility; 5-24: F1 generation formed by crossing durum wheat Jing DR691 and durum wheat Langdon. 2:3 Family lineage, of which 5-9: homozygous disease-resistant F1 cells 2:3 Family history, 10-19: F strains of immune system separation 2:3 Family history, 20-24: Homozygous F 2:3 Family lineage; white arrows indicate areas where genes can be traced. PmDR691 Specific bands. Amplification results show the markers. HENU1060 A 616 bp tightly linked specific band was amplified in the resistant parent durum wheat Jing DR691 and the resistant family, but no such band was amplified in the susceptible parent durum wheat Langdon and the susceptible family.

[0054] wheat powdery mildew resistance gene PmDR691 Derived from the durum wheat variety Jing DR691, this gene has not yet been reported to be located or cloned using map-based methods. The molecular markers provided in this invention... HENU1060 Detecting large populations for genetic mapping is helpful for gene sequencing. PmDR691 Fine-grained localization and map-based cloning are crucial for gene sequencing. PmDR691 The in-depth analysis of the efficient breeding and disease resistance mechanisms is of great significance.

[0055] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. Related to wheat powdery mildew resistance genes PmDR691 Closely linked molecular markers, characterized by: The molecular marker is located on wheat chromosome 7BL, at 696934274bp.

2. A primer set for identifying molecular markers, characterized in that: The primer set includes an upstream primer. HENU1060-F and downstream primers HENU1060-R upstream primer HENU1060-F The nucleotide sequence is shown in SEQ ID No. 1; the downstream primer HENU1060-R The nucleotide sequence is shown in SEQ ID No.

2.

3. A kit comprising the primer set of claim 2, characterized in that: It also includes testing reagents.

4. The primer set described in claim 2 for identifying wheat powdery mildew resistance genes. PmDR691 Applications in [the field].

5. The primer set described in claim 2 for map-based cloning identification of wheat powdery mildew resistance genes. PmDR691 Applications in [the field].

6. The application of the primer set according to claim 2 in the cultivation of powdery mildew resistant wheat.

7. A method for identifying wheat powdery mildew resistance genes PmDR691 The method is characterized by, Includes the following steps: (1) Extract genomic DNA from the wheat sample to be tested; (2) Using the primer set described in claim 2 as primers and the genomic DNA from step (1) as template, perform PCR amplification to obtain the amplification product; (3) The amplification product of step (2) is tested to determine whether it contains the wheat powdery mildew resistance gene. PmDR691 .

8. The method for identifying wheat powdery mildew resistance genes according to claim 7. PmDR691 The method is characterized by, The PCR amplification system in step (2) is as follows: 1.0 μL of 50 ng / μL wheat genomic DNA, 4 μL of PCR MasterMix, 0.5 μL of 5 μM upstream primer, 0.5 μL of 5 μM downstream primer, and 4 μL of sterile deionized water.

9. The method for identifying wheat powdery mildew resistance genes according to claim 7. PmDR691 The method is characterized by, The PCR amplification procedure in step (2) is as follows: pre-denaturation at 94℃ for 5 min; denaturation at 94℃ for 15 s, annealing at 60℃ for 15 s, extension for 40 s, 34 cycles; extension at 72℃ for 10 min; storage at 12℃.

10. The method for identifying wheat powdery mildew resistance genes according to any one of claims 7-9 PmDR691 The method is characterized by: If the amplification product is a specific band of 616 bp, it indicates that the wheat sample being tested contains a gene for resistance to powdery mildew. PmDR691 Conversely, the wheat sample being tested did not contain the wheat powdery mildew resistance gene. PmDR691 .

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