Molecular marker primer pair for aegilops ovalis 7M chromosome and application of molecular marker primer pair

Abstract

The invention discloses an aegilops ovalis 7M chromosome molecular marker primer pair. Comprising a primer pair 1, a primer pair 2, a primer pair 3, a primer pair 6, a primer pair 7, a primer pair 10, a primer pair 11, a primer pair 15, a primer pair 16, a primer pair 17, a primer pair 18, a primer pair 19, a primer pair 20, a primer pair 21, a primer pair 22, a primer pair 25, a primer pair 36, a primer pair 37, a primer pair 43, a primer pair 45, a primer pair 46, a primer pair 51, a primer pair 53, a primer pair 54, a primer pair 55, a primer pair 56, a primer pair 57 and a primer pair 59. The molecular marker primer pair is used for detecting the aegilops ovalis 7M chromosome or fragments thereof, the specific molecular marker of the 7M chromosome is developed, the specific molecular marker can be used for rapid detection of the aegilops ovalis 7M chromosome (or fragments), and a foundation is laid for further utilization of excellent aegilops ovalis genes in wheat genetic breeding.

Description

Technical Field

[0001] The invention relates to the field of biotechnology, in particular to a pair of molecular marker primers for chromosome 7M of Aegilops ovalis and application thereof. Background Art

[0002] Wheat chromosome engineering technology is to selectively introduce chromosome sets, chromosomes or chromosome fragments into common wheat. This allows wheat breeders to purposefully improve the stress resistance, yield and quality of wheat, thereby accelerating the wheat breeding process. Regardless of the form in which exogenous chromosomes are transferred to common wheat, these materials have more or less adverse effects on wheat breeding. Almost all created materials cannot be directly used in the actual production of wheat. However, they can be used as a very valuable special basic material to create translocation lines or introgression line materials that are more conducive to breeding. Therefore, it is particularly important to identify the created exogenous chromosomes. The methods mainly include morphological markers, cytological observations, in situ hybridization analysis, molecular marker detection, SLAF-seq sequencing, 660K chips and other technical means. Among them, the use of molecular marker detection is the fastest and lowest cost method.

[0003] Aegilops geniculata Roth (2n=4x=28) is an annual self-pollinating allotetraploid species of the genus Capra with a chromosome configuration of UgUgMgMg, in which the chromosome M group is derived from Ae. comosa Sm. (Molnar et al. 2013; Schneider et al. 2010), and the chromosome U group is derived from Ae. umbellulata Zhuk. It has a large number of genes that contribute to crop quality and resistance that modern common wheat does not have. It is a valuable gene resource library to be developed. Aegilops geniculata Roth contains rich disease resistance genes: resistance to stripe rust, leaf rust, powdery mildew and other excellent genes for high quality.

[0004] So far, the resistance genes introduced from Aegilops ovalis into common wheat are very limited, including the powdery mildew resistance gene Pm29, rust resistance genes Yr40, Lr57 and Sr53, and insect resistance genes on 1Mg and 7Mg. The excellent target genes of Aegilops ovalis can be transferred to common wheat through molecular breeding methods to cultivate new wheat lines (species) with excellent genes of Aegilops ovalis, which plays an important role in wheat molecular breeding. Although people have long recognized the great potential of wild species to improve wheat, the genetic diversity of wild species has not yet been effectively developed and utilized. Summary of the invention

[0005] The technical problem to be solved by the invention is to provide a pair of molecular marker primers for chromosome 7M of Aegilops ovalis and application thereof.

[0006] In order to solve the above technical problems, the technical solutions adopted by the present invention are as follows.

[0007] A molecular marker primer pair for Aegilops ovata 7M chromosome, comprising primer pair 1, primer pair 2, primer pair 3, primer pair 6, primer pair 7, primer pair 10, primer pair 11, primer pair 15, primer pair 16, primer pair 17, primer pair 18, primer pair 19, primer pair 20, primer pair 21, primer pair 22, primer pair 25, primer pair 36, primer pair 37, primer pair 43, primer pair 45, primer pair 46, primer pair 51, primer pair 53, primer pair 54, primer pair 55, primer pair 56, primer pair 57, and primer pair 59, wherein the molecular marker primer pair is used for detecting Aegilops ovata 7M chromosome or a fragment thereof;

[0008] The nucleotide sequences of primer pair 1 are shown in SEQ ID NO.1 and SEQ ID NO.2;

[0009] The nucleotide sequences of primer pair 2 are shown in SEQ ID NO.3 and SEQ ID NO.4;

[0010] The nucleotide sequences of primer pair 3 are shown in SEQ ID NO.5 and SEQ ID NO.6;

[0011] The nucleotide sequences of primer pair 6 are shown in SEQ ID NO.7 and SEQ ID NO.8;

[0012] The nucleotide sequences of primer pair 7 are shown in SEQ ID NO.9 and SEQ ID NO.10;

[0013] The nucleotide sequences of primer pair 10 are shown in SEQ ID NO.11 and SEQ ID NO.12;

[0014] The nucleotide sequences of primer pair 11 are shown in SEQ ID NO. 13 and SEQ ID NO. 14;

[0015] The nucleotide sequences of primer pair 15 are shown in SEQ ID NO. 15 and SEQ ID NO. 16;

[0016] The nucleotide sequences of primer pair 16 are shown in SEQ ID NO. 17 and SEQ ID NO. 18;

[0017] The nucleotide sequences of primer pair 17 are shown in SEQ ID NO. 19 and SEQ ID NO. 20;

[0018] The nucleotide sequences of primer pair 18 are shown in SEQ ID NO. 21 and SEQ ID NO. 22;

[0019] The nucleotide sequences of primer pair 19 are shown in SEQ ID NO. 23 and SEQ ID NO. 24;

[0020] The nucleotide sequences of primer pair 20 are shown in SEQ ID NO. 25 and SEQ ID NO. 26;

[0021] The nucleotide sequences of primer pair 21 are shown in SEQ ID NO. 27 and SEQ ID NO. 28;

[0022] The nucleotide sequences of primer pair 22 are shown in SEQ ID NO. 29 and SEQ ID NO. 30;

[0023] The nucleotide sequences of primer pair 25 are shown in SEQ ID NO.31 and SEQ ID NO.32;

[0024] The nucleotide sequences of primer pair 36 are shown in SEQ ID NO. 33 and SEQ ID NO. 34;

[0025] The nucleotide sequences of primer pair 37 are shown in SEQ ID NO. 35 and SEQ ID NO. 36;

[0026] The nucleotide sequences of primer pair 43 are shown in SEQ ID NO.37 and SEQ ID NO.38;

[0027] The nucleotide sequences of primer pair 45 are shown in SEQ ID NO.39 and SEQ ID NO.40;

[0028] The nucleotide sequences of primer pair 46 are shown in SEQ ID NO.41 and SEQ ID NO.42;

[0029] The nucleotide sequences of primer pair 51 are shown in SEQ ID NO. 43 and SEQ ID NO. 44;

[0030] The nucleotide sequences of primer pair 53 are shown in SEQ ID NO.45 and SEQ ID NO.46;

[0031] The nucleotide sequences of primer pair 54 are shown in SEQ ID NO. 47 and SEQ ID NO. 48;

[0032] The nucleotide sequences of primer pair 55 are shown in SEQ ID NO. 49 and SEQ ID NO. 50;

[0033] The nucleotide sequences of primer pair 56 are shown in SEQ ID NO.51 and SEQ ID NO.52;

[0034] The nucleotide sequences of primer pair 57 are shown in SEQ ID NO.53 and SEQ ID NO.54;

[0035] The nucleotide sequences of primer pair 59 are shown in SEQ ID NO.55 and SEQ ID NO.56.

[0036] Further preferably, the molar concentration ratio of the two primers in the primer pair is 1:1.

[0037] Application of a pair of molecular marker primers for chromosome 7M of Aegilops ovata in tracing the genetic stability of chromosome 7M of Aegilops ovata or its fragments.

[0038] The purpose of the molecular marker primer pair for the Aegilops ovata 7M chromosome is to screen, preliminarily screen, identify and assist in identifying wheat germplasm resources by detecting the Aegilops ovata 7M chromosome or its fragment.

[0039] Application of molecular marker primer pairs of chromosome 7M of Aegilops ovalis in identifying the amount of gene introduced into chromosome 7M of Aegilops ovalis.

[0040] A detection kit contains any one or any combination of Aegilops ovalis 7M chromosome molecular marker primer pairs and a PCR amplification reagent.

[0041] The present invention also includes a method for detecting exogenous chromosomes in the wheat genome, which comprises performing gene amplification and gel electrophoresis based on the primer pair described in claim 1, and detecting and identifying the exogenous chromosomes in the wheat genome based on the spectrum of the electrophoresis map.

[0042] Further preferably, the exogenous chromosome is: Aegilops ovalis 7M chromosome or a fragment thereof.

[0043] A method for designing chromosome-specific primers of Aegilops ovalis 7M comprises the following steps:

[0044] (1) Leaves of common wheat Chinese Spring, Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line were sent to BioMed Biotech for simplified genome sequencing to obtain the corresponding SLAF tags;

[0045] (2) The SLAF tags of Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line were compared with the SLAF tags obtained from Chinese Spring, and the sequences of the SLAF tags of Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line that had a sequence homology of ≥50% with the SLAF tag sequence obtained from Chinese Spring were eliminated;

[0046] (3) The SLAF tags obtained from the Chinese Spring-Aegilops ovata 7M addition line and the Chinese Spring-Aegilops ovata 7M substitution line obtained in the above step (2) were eliminated, and the SLAF tags obtained were compared with the SLAF tags of Aegilops ovata, and the sequences with similarity ≥ 80% were retained;

[0047] (4) comparing the SLAF tags retained by the Chinese spring-Aegilops ovata 7M addition line and the Chinese spring-Aegilops ovata 7M substitution line obtained in the above step (3), and retaining sequences with a sequence homology of ≥90%;

[0048] (5) The SLAF tag with a sequence homology of ≥90% finally obtained in step (4) is the chromosome-specific sequence of Aegilops ovalis 7M. n chromosomes of Aegilops ovalis 7M are randomly selected for primer design. The designed primers can amplify specific bands with a band size between 250-350bp in Aegilops ovalis SY159, Chinese Spring-Aegilops ovalis 7M addition line, and Chinese Spring-Aegilops ovalis 7M (7A) substitution line, but cannot amplify polymorphic bands in common wheat Chinese Spring.

[0049] The beneficial effect produced by adopting the above technical scheme is that: the present invention uses SLAF-seq technology to sequence common wheat Chinese Spring, Aegilops SY159, Chinese Spring-Aegilops 7M addition line and Chinese Spring-Aegilops 7M (7A) substitution line, and through sequence comparison, develops a specific molecular marker of the 7M chromosome, which can be used for the rapid detection of Aegilops 7M chromosome (or fragment). The molecular marker of Aegilops 7M chromosome developed by SLAF-seq sequencing in the present invention has not been reported yet, which lays a foundation for the further use of Aegilops 7M excellent genes in wheat genetic breeding. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 The electrophoresis results of the primer pairs SALF1, SALF2, SALF3, and SALF6 in different materials are shown in the figure, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line;

[0051] Figure 2 The electrophoresis results of the primer pairs SALF7, SALF10, SALF11, and SALF15 in different materials are shown in the figure, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line;

[0052] Figure 3 The electrophoresis results of amplification of primer pairs SALF16, SALF17, SALF18, and SALF19 in different materials are shown in the figure, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line;

[0053] Figure 4 The electrophoresis results of the primer pairs SALF20, SALF21, SALF22 and SALF25 amplified in different materials are shown in the figure, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line;

[0054] Figure 5 The electrophoresis results of amplification of primer pairs SALF36, SALF37, SALF43, and SALF45 in different materials are shown in the figure, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line;

[0055] Figure 6 The electrophoresis results of the primer pairs SALF46, SALF47, SALF53 and SALF54 amplified in different materials are shown in the figure, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line;

[0056] Figure 7 The graph shows the electrophoresis results of amplification of primer pairs SALF55, SALF56, SALF57, and SALF59 in different materials, where M: DL2000; 1: Chinese Spring; 2: Aegilops SY159; 3: Chinese Spring-Aegilops 7M addition line; 4: Chinese Spring-Aegilops 7M (7A) substitution line. DETAILED DESCRIPTION

[0057] The following examples illustrate the present invention in detail. The various raw materials and equipment used in the present invention are conventional commercial products and can be directly obtained by purchasing from the market. The experimental methods used in the following examples are conventional methods unless otherwise specified.

[0058] It should be understood that when used in the present specification and the appended claims, the term "comprising" indicates the presence of described features, wholes, steps, operations, elements and / or components, but does not exclude the presence or addition of one or more other features, wholes, steps, operations, elements, components and / or combinations thereof.

[0059] It should also be understood that the term “and / or” used in the specification and appended claims refers to any and all possible combinations of one or more of the associated listed items, and includes these combinations.

[0060] References to "one embodiment" or "some embodiments" etc. described in the specification of this application mean that one or more embodiments of the present application include specific features, structures or characteristics described in conjunction with the embodiment. Therefore, the statements "in one embodiment", "in some embodiments", "in some other embodiments", "in some other embodiments", etc. that appear in different places in this specification do not necessarily refer to the same embodiment, but mean "one or more but not all embodiments", unless otherwise specifically emphasized in other ways. The terms "including", "comprising", "having" and their variations all mean "including but not limited to", unless otherwise specifically emphasized in other ways.

[0061] In addition, in the description of the present application specification and the appended claims, the terms "first", "second", "third", etc. are only used to distinguish the descriptions and cannot be understood as indicating or implying relative importance.

[0062] The following will be combined with specific embodiments of the present invention to clearly and completely describe the technical solution of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0063] Example 1: Screening of Aegilops ovalis 7M chromosome-specific sequences

[0064] Experimental materials: common wheat Chinese spring (T. aestivum, 2n=6x=42), Aegilops ovalis SY159, Chinese spring-Aegilops ovalis 7M addition line, Chinese spring-Aegilops ovalis 7M (7A) substitution line.

[0065] Methods: Leaves of common wheat Chinese Spring, Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M (7A) substitution line were sent to BioMed Biotech for simplified genome sequencing, and the obtained SLAF tags were screened according to the following steps:

[0066] (1) The SLAF tags of Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line were compared with the SLAF tags obtained from Chinese Spring, and the sequences of the SLAF tags of Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line that had a sequence homology of ≥50% with the SLAF tag sequence obtained from Chinese Spring were eliminated;

[0067] (2) The SLAF tags obtained from the Chinese Spring-Aegilops ovata 7M addition line and the Chinese Spring-Aegilops ovata 7M substitution line obtained in the above step (1) were eliminated, and the SLAF tags obtained were compared with the SLAF tags of Aegilops ovata, and the sequences with similarity ≥ 80% were retained;

[0068] (3) comparing the SLAF tags retained by the Chinese spring-Aegilops ovale 7M addition line and the Chinese spring-Aegilops ovale 7M substitution line obtained in the above step (2), and retaining sequences with a sequence homology of ≥90%;

[0069] The SLAF tags finally obtained were the chromosome 7M specific sequences of Aegilops ovalis, totaling 339.

[0070] 60 of the 339 specific sequences obtained above were randomly selected for primer design. The designed primers can amplify specific bands (band size is about 300 bp) in Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M (7A) substitution line, but cannot amplify polymorphic bands in common wheat Chinese Spring. These primers are 7M chromosome-specific primers. The marker screening results are shown in Table 1.

[0071] Table 1 shows the detection results of 28 SALF molecular markers in various wheat

[0072] Primer number Chinese Spring SY159 7M Additional System 7M(7A) Replacement Series SALF1 - + + + SALF2 - + + + SALF3 - + + + SALF6 - + + + SALF7 - + + + SALF10 - + + + SALF11 - + + + SALF15 - + + + SALF16 - + + + SALF17 - + + + SALF18 - + + + SALF19 - + + + SALF20 - + + + SALF21 - + + + SALF22 - + + + SALF25 - + + + SALF36 - + + + SALF37 - + + + SALF43 - + + + SALF45 - + + + SALF46 - + + + SALF51 - + + + SALF53 - + + + SALF54 - + + + SALF55 - + + + SALF56 - + + + SALF57 - + + + SALF59 - + + +

[0073] Note: “-” represents the size is different from that of Aegilops SY159, and “+” represents the size is the same as that of Aegilops SY159.

[0074] Example 2: PCR amplification

[0075] PCR reaction system: For a 10 μl sample amplification system, the following reagents are required: 10× Taq Buffer (containing Mg 2+ ) 1.0 μl, dNTPs (2.5 mmol / L) 0.8 μl, upstream and downstream primers 0.5 μl each (concentration 0.5 μmol / L), Taq DNA Polymerase (2.5 U / μl) 0.12 μl, DNA template 1 μl (concentration 200 ng / μl) and ddH2O 6.08 μl.

[0076] PCR amplification procedure: First, perform a pre-denaturation reaction at 94°C for 3 minutes; then perform 34 cycles, each cycle including the following three steps: denaturation at 94°C for 30 seconds, annealing temperature of 54-64°C (45 seconds) and extension at 72°C for 1 minute. Then perform a final extension reaction at 72°C for 10 minutes. Finally, after the sample amplification is completed, the reaction system is stored at 4°C.

[0077] The PCR products were detected by 2% agarose gel electrophoresis.

[0078] After PCR amplification and verification in common wheat Chinese Spring, Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M (7A) substitution line, 28 pairs of 7M chromosome-specific primers were obtained, the molar concentration ratio of the two primers in the primer pair was 1:1, and the molecular marker development efficiency reached 46.7%. Among them, the primer sequences of the 28 pairs of specific molecular markers are shown in Table 1.

[0079] Table 2 DNA sequences of 28 molecular marker primer pairs screened

[0080]

[0081]

[0082] Example 3: Verification

[0083] like Figure 1-7 As shown, they are respectively amplification electrophoresis diagrams of 28 pairs of molecular marker primers in common wheat Chinese spring, Aegilops ovatus SY159, common wheat-Aegilops ovatus 7M addition line and common wheat-Aegilops ovatus 7M (7A) substitution line. It can be seen from the figures that the 28 pairs of primers can amplify specific bands in Aegilops ovatus SY159, Chinese spring-Aegilops ovatus 7M addition line and Chinese spring-Aegilops ovatus 7M (7A) substitution line, but can not amplify polymorphic bands in common wheat Chinese spring, indicating that the specific molecular markers of Aegilops ovatus 7M chromosome developed by the present invention can be used for the rapid detection of Aegilops ovatus 7M chromosome (or fragment), which lays a foundation for the further utilization of Aegilops ovatus excellent genes in wheat genetic breeding.

[0084] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that numerous changes, modifications, substitutions and variations may be made to the examples without departing from the principles and spirit of the invention, the scope of the invention being defined by the appended claims and their equivalents.

[0085] In the above embodiments, the description of each embodiment has its own emphasis. For parts that are not described or recorded in detail in a certain embodiment, reference can be made to the relevant descriptions of other embodiments.

[0086] The embodiments described above are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. Such modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention, and should all be included in the protection scope of the present invention.

Claims

1. A pair of molecular marker primers for chromosome 7M of Aegilops ovalis, characterized in that: The invention comprises primer pair 1, primer pair 2, primer pair 3, primer pair 6, primer pair 7, primer pair 10, primer pair 11, primer pair 15, primer pair 16, primer pair 17, primer pair 18, primer pair 19, primer pair 20, primer pair 21, primer pair 22, primer pair 25, primer pair 36, primer pair 37, primer pair 43, primer pair 45, primer pair 46, primer pair 51, primer pair 53, primer pair 54, primer pair 55, primer pair 56, primer pair 57 and primer pair 59, wherein the molecular marker primer pair is used for detecting chromosome 7M of Aegilops ovalis or a fragment thereof; The nucleotide sequences of primer pair 1 are shown in SEQ ID NO. 1 and SEQ ID NO. 2; The nucleotide sequences of primer pair 2 are shown in SEQ ID NO. 3 and SEQ ID NO. 4; The nucleotide sequences of primer pair 3 are shown in SEQ ID NO. 5 and SEQ ID NO. 6; The nucleotide sequences of primer pair 6 are shown in SEQ ID NO. 7 and SEQ ID NO. 8; The nucleotide sequences of primer pair 7 are shown in SEQ ID NO. 9 and SEQ ID NO. 10; The nucleotide sequences of primer pair 10 are shown in SEQ ID NO. 11 and SEQ ID NO. 12; The nucleotide sequences of primer pair 11 are shown in SEQ ID NO. 13 and SEQ ID NO. 14; The nucleotide sequences of primer pair 15 are shown in SEQ ID NO. 15 and SEQ ID NO. 16; The nucleotide sequences of primer pair 16 are shown in SEQ ID NO. 17 and SEQ ID NO. 18; The nucleotide sequences of primer pair 17 are shown in SEQ ID NO. 19 and SEQ ID NO. 20; The nucleotide sequences of primer pair 18 are shown in SEQ ID NO. 21 and SEQ ID NO. 22; The nucleotide sequences of primer pair 19 are shown in SEQ ID NO. 23 and SEQ ID NO. 24; The nucleotide sequences of primer pair 20 are shown in SEQ ID NO. 25 and SEQ ID NO. 26; The nucleotide sequences of primer pair 21 are shown in SEQ ID NO. 27 and SEQ ID NO. 28; The nucleotide sequences of primer pair 22 are shown in SEQ ID NO. 29 and SEQ ID NO. 30; The nucleotide sequences of primer pair 25 are shown in SEQ ID NO. 31 and SEQ ID NO. 32; The nucleotide sequences of primer pair 36 are shown in SEQ ID NO. 33 and SEQ ID NO. 34; The nucleotide sequences of primer pair 37 are shown in SEQ ID NO. 35 and SEQ ID NO. 36; The nucleotide sequences of primer pair 43 are shown in SEQ ID NO. 37 and SEQ ID NO. 38; The nucleotide sequences of primer pair 45 are shown in SEQ ID NO. 39 and SEQ ID NO. 40; The nucleotide sequences of primer pair 46 are shown in SEQ ID NO. 41 and SEQ ID NO. 42; The nucleotide sequences of primer pair 51 are shown in SEQ ID NO. 43 and SEQ ID NO. 44; The nucleotide sequences of primer pair 53 are shown in SEQ ID NO. 45 and SEQ ID NO. 46; The nucleotide sequences of primer pair 54 are shown in SEQ ID NO. 47 and SEQ ID NO. 48; The nucleotide sequences of primer pair 55 are shown in SEQ ID NO. 49 and SEQ ID NO. 50; The nucleotide sequences of primer pair 56 are shown in SEQ ID NO. 51 and SEQ ID NO. 52; The nucleotide sequences of primer pair 57 are shown in SEQ ID NO. 53 and SEQ ID NO. 54; The nucleotide sequences of primer pair 59 are shown in SEQ ID NO. 55 and SEQ ID NO.

56.

2. The molecular marker primer pair according to claim 1, wherein the molar concentration ratio of the two primers in the primer pair is 1:

1.

3. Use of the molecular marker primer pair according to claim 1 in identifying whether wheat plants have Aegilops ovalis 7M chromosome or its fragment.

4. Use of the molecular marker primer pair according to claim 1 in tracing the genetic stability of Aegilops ovalis 7M chromosome or its fragments.

5. Use of the molecular marker primer pair according to claim 1, for screening, preliminary screening, identification, and auxiliary identification of wheat germplasm resources by detecting Aegilops ovalis 7M chromosome or its fragment.

6. Use of the molecular marker primer pair according to claim 1 in identifying the amount of gene introduced into chromosome 7M of Aegilops ovalis.

7. A detection kit, characterized in that: Contains any one or any combination of the primer pairs described in claim 1, and a PCR amplification reagent.

8. A method for detecting exogenous chromosomes in a wheat genome, characterized in that: Gene amplification and gel electrophoresis are performed based on the primer pair described in claim 1, and the exogenous chromosomes in the wheat genome are detected and identified based on the spectrum of the electrophoresis map.

9. The method according to claim 8, characterized in that: The exogenous chromosome is: Aegilops ovalis 7M chromosome or a fragment thereof.

10. A method for designing chromosome-specific primers for Aegilops ovalis 7M, characterized in that: The following steps are involved: (1) Leaves of common wheat Chinese Spring, Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line were sent to BioMed Biotech for simplified genome sequencing to obtain the corresponding SLAF tags; (2) The SLAF tags of Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line were compared with the SLAF tags obtained from Chinese Spring, and the sequences in the SLAF tags of Aegilops SY159, Chinese Spring-Aegilops 7M addition line, and Chinese Spring-Aegilops 7M substitution line that had a sequence homology of ≥50% with the SLAF tag sequence obtained from Chinese Spring were removed; (3) The SLAF tags obtained from the Chinese Spring-Aegilops ovata 7M addition line and the Chinese Spring-Aegilops ovata 7M substitution line obtained in the above step (2) were removed, and the SLAF tags obtained from the Chinese Spring were compared with the SLAF tags of Aegilops ovata, and the sequences with similarity ≥ 80% were retained; (4) comparing the SLAF tags retained by the Chinese spring-Aegilops ovata 7M addition line and the Chinese spring-Aegilops ovata 7M substitution line obtained in step (3) above, and retaining sequences with a sequence homology of ≥ 90%; (5) The SLAF tag with a sequence homology of ≥90% finally obtained in step (4) is the specific sequence of the chromosome of Aegilops ovalis 7M. n chromosomes of Aegilops ovalis 7M are randomly selected for primer design. The designed primers can amplify specific bands with a band size between 250-350 bp in Aegilops ovalis SY159, Chinese Spring-Aegilops ovalis 7M addition line, and Chinese Spring-Aegilops ovalis 7M (7A) substitution line, but cannot amplify polymorphic bands in common wheat Chinese Spring.

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