DCAPS molecular marker related to soybean internode spacing and application of dCAPS molecular marker

Abstract

The invention discloses a dCAPS molecular marker related to soybean internode spacing and application thereof, relates to the technical field of biology, and in particular relates to a dCAPS molecular marker related to soybean internode spacing and application thereof. According to the invention, the substance for detecting the polymorphism or genotype of the SNP site in the soybean genome can be used for identifying or assisting in identifying the average internode spacing character of the soybean and breeding the soybean; wherein the SNP site is deoxyribonucleotide at the 39828757 site of the 13th chromosome of the soybean genome, the version number of the reference genome is Glycine max Wm82. A2. V1, the SNP site corresponds to nucleotide at the 221 site of SEQ ID NO: 4, and the polymorphism of the SNP site is G or A. The invention further discloses a kit for detecting the soybean genome. The genotype GG homozygous soybean average internode spacing of the SNP is longer or candidate longer than the genotype AA homozygous soybean.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and in particular to a dCAPS molecular marker related to internode spacing in soybean and its applications. Background Technology

[0002] Soybean (Glycine max (Linn.) Merr.) is an important food crop in China with a cultivation history of 5,000 years. Historically known as "shu," its main production area is concentrated in Northeast China. Its seeds are rich in plant protein, containing 35%–45%, making it an important source of high-quality protein. As a high-protein, high-fat, and high-energy crop, soybeans have an oil content of approximately 17%–25% and are widely used. They are commonly processed into various soy products, extracted for soybean oil, brewed for soy sauce, or used for protein extraction, occupying a key position in the global agricultural product landscape. Statistics show that soybeans account for 61% of global oilseed crop production, 70% of protein powder production, and 61% of edible oil production. In 2021, approximately 120.5 million hectares of arable land worldwide yielded nearly 371.7 million tons of soybeans, providing not only a large amount of edible oil and protein for humans but also serving as a core protein source for animal feed. However, soybean yields are far lower than those of major food crops such as rice and wheat. The latter two crops have been bred into semi-dwarf varieties through plant dwarfing techniques, effectively reducing the risk of lodging and enabling increased yields under high-density planting. The yield formation mechanism of soybeans differs from that of soybeans. Soybean plant height is determined by both the number of nodes on the main stem and the length of the internodes, with the number of nodes directly affecting the number of pods and thus yield. Therefore, shortening the internode length and achieving plant dwarfing without reducing the number of nodes on the main stem is a potential key pathway to increase soybean yield.

[0003] The development of molecular marker technology has provided new technical means for improving internode spacing in soybeans. From early RFLP and RAPD markers to PCR-based SSR markers, and then to high-throughput SNP markers, molecular marker technology has been continuously evolving. Commonly used molecular markers in recent years include: (1) Simple sequence repeats (SSR), (2) Cleaved amplified polymorphic sequences (CAPS), and (3) Single nucleotide polymorphisms (SNP).

[0004] Recent studies have shown that the regulation of internode spacing in soybean involves multiple molecular networks, including gibberellin (GA) metabolism, light signal transduction, and transcriptional regulation. For example, DWARF MUTANT 1 (DW1) encodes an enantiomeric kauriene synthase, a key enzyme in the gibberellin synthesis pathway, which regulates interstitial cell elongation and consequently, plant height and internode spacing in soybean. The mutant dw1 shows reduced GA bioactivity, leading to dwarfing. Overexpression of the GA2ox8B gene can reduce bioactive GA content, decrease internode length, and inhibit apical elongation. The discovery of key genes such as rin1 and GmSPL3 provides an important foundation for the development of molecular markers for internode spacing. Furthermore, QTL mapping has identified several quantitative trait loci associated with internode spacing, such as qSI13-1 and qPH18. Most markers associated with internode spacing are neutral, indirectly linked to target genes only through linkage disequilibrium, and do not directly act on functional sites regulating the trait. Furthermore, this association is easily affected by genetic background and environmental conditions. Soybean internode spacing is a quantitative trait controlled by multiple genes, regulated synergistically by major and minor genes.

[0005] In conclusion, the discovery and development of molecular markers that regulate internode spacing in soybean and their application in modern breeding practices are of great scientific significance and practical value for my country to breed soybean varieties suitable for dense planting and with strong lodging resistance, thereby breaking through yield bottlenecks. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to provide a dCAPS molecular marker related to soybean internode spacing and its application.

[0007] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is as follows.

[0008] The application of substances for detecting SNP polymorphisms or genotypes in the soybean genome in any of the following situations.

[0009] (1) To identify or assist in the identification of the average internode spacing trait in soybeans;

[0010] (2) Screening or breeding soybean single plants, lines, varieties or strains with long average internode spacing;

[0011] (3) Screening or breeding soybean single plants, lines, varieties or strains with short average internode spacing.

[0012] (4) Soybean breeding;

[0013] (5) Prepare products for identification or auxiliary identification of the average internode spacing trait of soybeans;

[0014] (6) Prepare or select soybean single plants, lines, strains or varieties with long average internode spacing;

[0015] (7) Prepare or select soybean single plants, lines, varieties or strains with short average internode spacing;

[0016] (8) Prepare products for soybean breeding or soybean breeding assistance;

[0017] (9) Application in soybean plant architecture improvement;

[0018] (10) Application in the preparation of soybean plant type improvement products;

[0019] The SNP site is the deoxyribonucleotide at position 39828757 on chromosome 13 of the soybean genome, reference genome version number: Glycine max Wm82.a2.v1. The SNP site corresponds to nucleotide 221 of SEQ ID NO: 4, and the SNP site polymorphism is G or A.

[0020] A method for identifying or assisting in the identification of the average internode spacing trait in soybean includes detecting the genotype of a SNP site in the soybean genome to be tested, and identifying or assisting in the identification of the average internode spacing trait in soybean based on the genotype. The SNP site is the deoxyribonucleotide at position 39828757 on chromosome 13 of the soybean genome, reference genome version number: Glycine maxWm82.a2.v1, the SNP site corresponds to nucleotide 221 of SEQ ID NO: 4, and the SNP site polymorphism is G or A.

[0021] More preferably, the method for detecting the genotype of the soybean to be tested includes the following steps:

[0022] Using the genomic DNA of the soybean to be tested as a template, PCR amplification was performed using the primer pair shown in SEQ ID NO:1 and SEQ ID NO:2 to obtain PCR products;

[0023] The PCR product was digested with the restriction endonuclease NcoI;

[0024] Genotype can be determined by detecting the size of the enzyme digestion product bands via electrophoresis.

[0025] If the band sizes of the enzyme digestion products are 219 bp and 29 bp, then the soybean genotype to be tested is homozygous for the A allele.

[0026] If the enzyme digestion product band size is 248 bp, then the soybean genotype to be tested is homozygous for the G allele.

[0027] More preferably, the genotype of the SNP is GG or AA, where the GG genotype is a homozygous form of the SNP being G, and the AA genotype is a homozygous form of the SNP being A; the identification or auxiliary identification of the average internode distance trait of soybean based on the SNP genotype specifically involves: the average internode distance of soybeans homozygous for the GG genotype is longer than or can be longer than that of soybeans homozygous for the AA genotype.

[0028] The application of the above methods in soybean breeding.

[0029] The method for soybean breeding, wherein the method is M1 or M2,

[0030] M1. The method includes detecting the genotype of the SNP in claim 1 in the soybean genome, selecting soybeans with the genotype AA of the SNP as parents for breeding, wherein AA is a homozygous type of the SNP being A, and the breeding objective of the method includes selecting soybeans with a short average internode distance.

[0031] M2. The method includes detecting the genotype of the SNP in claim 1 in the soybean genome, selecting soybeans with the genotype GG of the SNP as parents for breeding, wherein GG is a homozygous type of the SNP being G, and the breeding objective of the method includes selecting soybeans with a long average internode distance.

[0032] A product containing a substance for detecting polymorphisms or genotypes of SNP sites in the soybean genome, wherein the product is any one of the following:

[0033] C1) Products that detect single nucleotide polymorphisms or genotypes related to the average internode spacing trait in soybeans;

[0034] C2) Products used for identifying or assisting in the identification of the average internode spacing trait in soybeans;

[0035] C3) Products used in soybean breeding;

[0036] C4) Screening or breeding soybean single plants, lines, varieties or strains with long average internode spacing;

[0037] C5) Screening or breeding of soybean single plants, lines, strains, or varieties with short average internode spacing.

[0038] In the above applications or products, the substance is as follows:

[0039] D1), D2), or D3):

[0040] D1) Contains a primer composition that amplifies soybean genomic DNA fragments including the SNP sites;

[0041] D2) PCR reagents containing the primer composition described in D1);

[0042] D3) A kit containing the primer composition described in D1) or the PCR reagent described in D2);

[0043] The primer composition consists of primer dCAPS-F and primer dCAPS-R;

[0044] The primer dCAPS-F is a single-stranded DNA molecule whose nucleotide sequence is SEQ ID NO: 1;

[0045] The primer dCAPS-R is a single-stranded DNA molecule whose nucleotide sequence is SEQ ID NO: 2.

[0046] DNA molecule, nucleotide sequence as shown in SEQ ID NO: 4.

[0047] The above-mentioned DNA molecules are used in any of the following applications:

[0048] (1) To identify or assist in the identification of the average internode spacing trait in soybeans;

[0049] (2) Screening or breeding soybean single plants, lines, varieties or strains with long average internode spacing;

[0050] (3) Screening or breeding soybean single plants, lines, varieties or strains with short average internode spacing;

[0051] (4) Soybean breeding;

[0052] (5) Prepare products for identification or auxiliary identification of the average internode spacing trait of soybeans;

[0053] (6) Prepare or select soybean single plants, lines, strains or varieties with long average internode spacing;

[0054] (7) Prepare or select soybean single plants, lines, varieties or strains with short average internode spacing;

[0055] (8) Prepare soybean breeding products.

[0056] The beneficial effects of adopting the above technical solution are as follows:

[0057] Compared with existing technologies, the present invention has the following advantages:

[0058] (1) This invention provides a SNP that is significantly associated with the internode spacing trait in soybean. Validation in a recombinant inbred line (RIL) population constructed from Williams82 (W82) (long internode, GG) and GDL025 (ant egg) (short internode, AA) showed that the dCAPS-1 marker was highly significantly associated with the average internode spacing trait in soybean (P<0.0001). The average internode spacing of the material carrying the GG genotype was significantly greater than that of the material carrying the AA genotype. This provides a direct basis for early and accurate screening of individual plants with the ideal short internode phenotype using molecular markers, effectively overcoming the limitations of traditional breeding that relies on later field phenotypic measurements, significantly shortening the breeding cycle, and improving selection efficiency.

[0059] (2) The marker developed in this invention can be directly used for marker-assisted selection. Through this marker, breeders can quickly identify and screen individuals carrying the favorable allele (A, short internode) during the seedling stage, thereby purposefully introducing the superior traits of short internode, dwarfism, and lodging resistance into high-yield or high-quality soybean varieties, accelerating the breeding of new varieties suitable for dense planting and with strong lodging resistance, and providing key technical support for breaking through the bottleneck of soybean yield. Attached Figure Description

[0060] Figure 1 This is a frequency distribution diagram of intersegment length in the RIL population in Embodiment 2 of the present invention;

[0061] Figure 2 Phenotypic diagrams of W82 and GDL025 in Embodiment 1 of the present invention;

[0062] Figure 3 This is a phenotypic difference diagram of the average pitch of W82 and GDL025 in Embodiment 1 of the present invention;

[0063] Figure 4 This is a gel image showing the parental differences in molecular markers in Example 2 of the present invention;

[0064] Figure 5 This is a gel image showing the differences in molecular markers among the progeny RIL population in Example 2 of this invention.

[0065] Figure 6 This is a statistical analysis diagram of molecular markers in the progeny RIL population in Example 2 of the present invention;

[0066] Figure 7 These are field survey photos from Embodiment 1 of the present invention. Detailed Implementation

[0067] The following embodiments illustrate the present invention in detail. All raw materials and equipment used in the present invention are commercially available products and can be directly obtained through market purchase. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods.

[0068] It should be understood that, when used in this application specification and the appended claims, the term "comprising" indicates the presence of the described features, integrals, steps, operations, elements and / or components, but does not exclude the presence or addition of one or more other features, integrals, steps, operations, elements, components and / or a collection thereof.

[0069] It should also be understood that the term “and / or” as used in this application specification and the appended claims means any combination of one or more of the associated listed items and all possible combinations, and includes such combinations.

[0070] References to "one embodiment" or "some embodiments" as described in this application specification mean that one or more embodiments of this application include the features described in connection with that embodiment.

[0071] Defined features, structure, or characteristics. Therefore, the phrases "in one embodiment," "in some embodiments," "in other embodiments," "in still other embodiments," etc., appearing in different places in this specification do not necessarily refer to the same embodiment, but rather mean "one or more, but not all, embodiments," unless otherwise specifically emphasized. The terms "comprising," "including," "having," and variations thereof mean "including, but not limited to," unless otherwise specifically emphasized.

[0072] Furthermore, in the description of this application and the appended claims, the terms "first," "second," "third," etc., are used only to distinguish descriptions and should not be construed as indicating or implying relative importance.

[0073] The technical solution of the present invention will be clearly and completely described below with reference to specific embodiments. 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 skilled in the art without creative effort are within the scope of protection of the present invention.

[0074] Example 1, Materials and Methods

[0075] 1.1 Test Materials

[0076] This experiment constructed a new RIL population by crossing Williams82 (W82), a RIL with longer internode length, and GDL025 (Ant Egg), a RIL with shorter internode length. This RIL population contained 151 families. The specific phenotypes and mean internode length differences between Williams82 (W82) and GDL025 (Ant Egg) are shown in [link to relevant documentation]. Figure 2 and Figure 3 .

[0077] 1.2 Test Methods

[0078] The parent lines and 151 RILs were planted in June 2024 at the Mazhuang Experimental Station in Gaocheng District, Hebei Province. Figure 7 (37.51°N, 114.39°E), randomized block design, with row spacing of 50 cm and plant spacing of 15 cm for each family. Conventional management measures were adopted in the field. After the plants matured, the middle 3 single plants of each family were randomly selected for harvesting, and the internode length was measured in centimeters, accurate to 0.1 cm.

[0079] 1.3 Statistics and Analysis

[0080] SPSS 27.0 was used to perform maximum, minimum, coefficient of variation, skewness, kurtosis and normality analysis on the obtained phenotypic data, and analysis of variance was performed on haplotype phenotypic differences.

[0081] 1.4 QTL Positioning

[0082] Based on the constructed high-density genetic linkage map, QTLs were detected using the interval mapping (IM) method in the Mapping QTL6 software, combined with population phenotypic data. A LOD greater than 2.5 was used as the presence threshold. The additive effect and phenotypic contribution rate of each QTL were calculated based on the software results. A positive additive effect indicates that the enhancing allele originates from the paternal parent, while a negative additive effect indicates that it originates from the maternal parent. The QTL naming principle is 'q' + the abbreviation of the target trait + chromosome or linkage group band.

[0083] 1.5 Molecular Marker Development

[0084] Based on the initial localization, according to the soybean genome sequencing data of this RIL population, a polymorphism of G / A was found at position 39828757 on chromosome 13 (reference genome version number: Glycine max Wm82 .a2 .v1), which is associated with soybean internode length.

[0085] The restriction endonuclease NocⅠ was selected using online enzyme recognition software (website: http: / / indcaps.kieber.cloudapps.unc.edu / results), and primer sequences were designed. The underlined bases in primer dCAPS-R are artificially introduced mismatched bases. When the base at the mutation site is G, the amplification product of primer pair dCAPS-F / dCAPS-R does not contain the NocⅠ recognition sequence and cannot be digested by NocⅠ. Conversely, when the base at the mutation site is A, the amplification product of primer pair dCAPS-F / dCAPS-R contains the NocⅠ recognition sequence and can be digested by NocⅠ.

[0086] Table 1 dCAPS-1 sites

[0087]

[0088] 1.6 Genotyping

[0089] 1.6.1 Test Materials

[0090] The test materials were derived from 52 accessions (F6 generation) of the RIL population. The RIL population was constructed by crossing Williams82 (W82, G genotype) with a longer internode length and GDL025 (ant egg, A genotype) with a shorter internode length. The RIL population was then self-crossed for 5 generations to achieve homozygous genotypes for each individual.

[0091] 1.6.2 DNA Extraction

[0092] The method for extracting soybean genomic DNA is as follows:

[0093] (1) Take about 100-500 mg of fresh soybean leaves into a 2 ml centrifuge tube, add three 3 mm steel balls, cool with liquid nitrogen and then grind quickly with a grinder to ensure that the sample is fully pulverized.

[0094] (2) Add 800 µL of crude extraction buffer to the sample centrifuge tube and vortex the sample for 1 minute to mix it thoroughly. Let it stand at room temperature for 10 min to allow the soybean leaf tissue to fully lyse.

[0095] (3) Centrifuge at 12000 rpm for 10 min, take 450 µL and put it into a new centrifuge tube, add 450 µL of pre-cooled isopropanol, shake vigorously, place in a -20℃ refrigerator to precipitate for 30 min, and then centrifuge at 12000 rpm for 10 min.

[0096] (5) After centrifugation, discard the liquid, add 1 ml of 75% ethanol, shake vigorously, then centrifuge at 12000 rpm for 5 min and discard the supernatant.

[0097] (6) Repeat step (5).

[0098] (7) After drying at 65℃ for 20 min, add 50 µL of ultrapure water.

[0099] (8) Quantitative detection of DNA was performed using an ultraviolet spectrophotometer. Qualified DNA was stored at -20 ℃ for later use.

[0100] 1.6.3 PCR Amplification

[0101] Configure the PCR reaction system and set up the PCR program for amplification:

[0102] Table 2 PCR reaction system

[0103]

[0104] Table 3 PCR amplification program

[0105]

[0106] After the PCR reaction is completed, 10 μL of the PCR product is subjected to 1% agarose gel electrophoresis. The results are then scanned using a gel imaging system. If the PCR amplification product contains only one main band with a fragment size of 248 bp, it indicates that the target fragment has been successfully amplified.

[0107] 1.6.4 Enzyme digestion

[0108] Enzyme digestion system:

[0109] Table 4 Enzyme digestion system

[0110]

[0111] Reaction conditions: 37℃ for 2 hours, followed by inactivation at 65℃ for 5 minutes.

[0112] 1.6.5 Electrophoresis: The enzyme digestion products were subjected to agarose gel electrophoresis. The specific steps are as follows:

[0113] (1) Electrophoresis buffer preparation: 1×TAE buffer (powder, Solarbio) added to 1 L of distilled water

[0114] (2) Preparation of 6% agarose gel: Weigh 6 g agarose and add 100 mL 1×TAE buffer. Heat in a microwave oven until completely dissolved. After the solution cools down, add 4 μL nucleic acid dye, pour into the gel tank, and insert two rows of 10 μL combs.

[0115] (3) Electrophoresis: Use a pipette to add 4 μL of 2000 DNA Marker to the first well, followed by 8 μL of the enzyme digestion product to the subsequent wells. Turn on the electrophoresis apparatus and set the electrophoresis parameters: constant voltage 120 volts, electrophoresis time approximately 2 hours. After electrophoresis, scan the results using a gel imaging system.

[0116] Because the amplification products of the GG genotype soybean material do not contain the restriction endonuclease NcoI recognition sequence, the digestion products cannot be digested by NcoI, and the band length of the digested products detected by electrophoresis is 248 bp. In contrast, the amplification products of the AA genotype soybean material contain the restriction endonuclease NcoI recognition sequence, so the digestion products can be digested by NcoI, and the band lengths of the digested products detected by electrophoresis are 219 bp and 29 bp, respectively. The GG genotype refers to the homozygous type where the deoxyribonucleotide at position 39828757 of chromosome 13 in the soybean genome is G, and the AA genotype refers to the homozygous type where the deoxyribonucleotide at position 39828757 of chromosome 13 in the soybean genome is A.

[0117] Example 2, Results

[0118] 2.1 Analysis of phenotypic genetic variation in mean internode spacing of the RIL population

[0119] Plant height is measured as the length from the cotyledon node to the apical meristem after the plant matures. Internode length is measured as follows: the distance between the cotyledon and the true leaf is the first internode length; the distance between the true leaf and the first trifoliate leaf is the second internode length; and so on. The average internode length of the same plant is taken as the average internode length.

[0120] The phenotypic identification results of the RIL population showed that the mean internode length varied from 1.90 to 5.2, with a coefficient of variation of 20.01%. The internode length phenotypic kurtosis of the RIL population was 0.261, and the skewness was 0.486, exhibiting an approximately normal distribution. Figure 1 ).

[0121] 3.2 QTL localization of RIL population

[0122] Based on the phenotypic and genotypic data of the average internode spacing of the RIL population, QTL analysis was performed on the average internode spacing of the RIL population planted at the Mazhuang experimental station in Gaocheng District, Hebei Province using the IM method of Mapping QTL6.0 software. The results showed that the internode spacing was located on chromosome 13, with the interval Gm_39752436-Gm_39884865, and the additive effect originated from the maternal parent (Table 5).

[0123] Table 5. QTL location results of average internode spacing in soybeans from the RIL population.

[0124]

[0125] 3.2 Genotype Identification Results

[0126] Based on the initial localization, dCAPS-1 markers linked to QTLs were developed using SNP variations among parents (Table 1). First, detection was performed between parents: dCAPS-1 markers showed differences between parents. Williams82 (W82) corresponded to the A banding type (GG genotype), and GDL025 (ant egg) corresponded to the B banding type (AA genotype). A gel permeation of parental differences in molecular markers is shown below. Figure 4 .

[0127] Molecular markers were detected in progeny families of field-grown RIL populations; differential glucograms of molecular markers in progeny RIL populations are shown in the following figures. Figure 5 Of these, 25 samples corresponded to strip type A, and 26 samples corresponded to strip type B. The average pitch difference between the two strip types was significant (P<0.0001). Figure 6 (Tables 6 and 7).

[0128] Table 6. Results of molecular marker and mean internode spacing detection in the progeny RIL population

[0129]

[0130] Table 7 Statistical analysis of the relationship between molecular markers and mean internode spacing in progeny RIL populations

[0131]

[0132] The mean internode spacing of GG and AA genotype families was 5.08 cm and 3.77 cm, respectively. The internode spacing phenotype was significantly different between soybean materials with genotype GG and AA (P < 0.0001). Therefore, the dCAPS marker of this invention can be used for breeding soybean germplasm with the mean internode spacing trait. Furthermore, this invention accurately genotypes soybean with the mean internode spacing trait, and this molecular marker can be applied to early, large-scale germplasm screening to assist in the molecular breeding of functional soybeans.

[0133] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these examples without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

[0134] In the above embodiments, the descriptions of each embodiment have different focuses. For parts that are not described in detail or recorded in a certain embodiment, please refer to the relevant descriptions of other embodiments.

[0135] The above-described embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention, and should all be included within the protection scope of the present invention.

Claims

1. The application of substances for detecting SNP polymorphisms or genotypes in the soybean genome in any of the following: (1) To identify or assist in the identification of the average internode spacing trait in soybeans; (2) Screening or breeding soybean single plants, lines, varieties or strains with long average internode spacing; (3) Screening or breeding soybean single plants, lines, varieties or strains with short average internode spacing. (4) Soybean breeding; (5) Prepare products for identification or auxiliary identification of the average internode spacing trait of soybeans; (6) Prepare or select soybean single plants, lines, strains or varieties with long average internode spacing; (7) Prepare or select soybean single plants, lines, varieties or strains with short average internode spacing; (8) Prepare products for soybean breeding or soybean breeding assistance; (9) Application in soybean plant architecture improvement; (10) Application in the preparation of soybean plant type improvement products; The SNP site is the deoxyribonucleotide at position 39828757 on chromosome 13 of the soybean genome, reference genome version number: Glycine max Wm82.a2.v1. The SNP site corresponds to nucleotide 221 of SEQ ID NO: 4, and the SNP site polymorphism is G or A.

2. A method for identifying or assisting in the identification of the average internode spacing trait in soybeans, characterized in that: This includes detecting the genotype of SNP sites in the soybean genome, and identifying or assisting in the identification of the average internode spacing trait in soybean based on the genotype. The SNP site is the deoxyribonucleotide at position 39,828,757 on chromosome 13 of the soybean genome, with reference genome version number: Glycine max Wm82.a2.v1. The SNP site corresponds to nucleotide 221 of SEQ ID NO: 4, and the SNP site polymorphism is G or A.

3. The method according to claim 2, characterized in that: The method for detecting the genotype of soybean to be tested includes the following steps: Using the genomic DNA of the soybean to be tested as a template, PCR amplification was performed using the primer pair shown in SEQ ID NO:1 and SEQ ID NO:2 to obtain PCR products; The PCR product was digested with the restriction endonuclease NcoI; Genotype can be determined by detecting the size of the enzyme digestion product bands via electrophoresis. If the band sizes of the enzyme digestion products are 219 bp and 29 bp, then the soybean genotype to be tested is homozygous for the A allele. If the enzyme digestion product band size is 248 bp, then the soybean genotype to be tested is the G allele homozygous type.

4. The application according to claim 1 or the method according to claim 2, characterized in that: The genotype of the SNP is GG or AA, where GG is the homozygous type of the SNP being G, and AA is the homozygous type of the SNP being A; the identification or auxiliary identification of the average internode distance trait of soybean based on the SNP genotype specifically involves: soybeans with a homozygous GG genotype have an average internode distance that is longer than or can be longer than soybeans with a homozygous AA genotype.

5. The application of the method according to any one of claims 2 to 4 in soybean breeding.

6. A method for soybean breeding, characterized by: The method is M1 or M2. M1. The method includes detecting the genotype of the SNP in claim 1 in the soybean genome, selecting soybeans with the genotype AA of the SNP as parents for breeding, wherein AA is a homozygous type of the SNP being A, and the breeding objective of the method includes selecting soybeans with a short average internode distance. M2. The method includes detecting the genotype of the SNP in claim 1 in the soybean genome, selecting soybeans with the genotype GG of the SNP as parents for breeding, wherein GG is a homozygous type of the SNP being G, and the breeding objective of the method includes selecting soybeans with a long average internode distance.

7. A product containing a substance for detecting polymorphisms or genotypes of SNP sites in the soybean genome, characterized in that: The product is any one of the following: C1) Products that detect single nucleotide polymorphisms or genotypes related to the average internode spacing trait in soybeans; C2) Products used for identifying or assisting in the identification of the average internode spacing trait in soybeans; C3) Products used in soybean breeding; C4) Screening or breeding soybean single plants, lines, varieties or strains with long average internode spacing; C5) Screening or breeding of soybean single plants, lines, strains, or varieties with short average internode spacing.

8. The application according to claim 1 or the product according to claim 7, characterized in that: The substance is as follows: D1), D2), or D3): D1) Contains a primer composition that amplifies soybean genomic DNA fragments including the SNP sites; D2) PCR reagents containing the primer composition described in D1); D3) A kit containing the primer composition described in D1) or the PCR reagent described in D2); The primer composition consists of primer dCAPS-F and primer dCAPS-R; The primer dCAPS-F is a single-stranded DNA molecule whose nucleotide sequence is SEQ ID NO: 1; The primer dCAPS-R is a single-stranded DNA molecule whose nucleotide sequence is SEQ ID NO:

2.

9. A DNA molecule, characterized by: The nucleotide sequence is shown in SEQ ID NO:

4.

10. The use of the DNA molecule of claim 9 in any of the following: (1) To identify or assist in the identification of the average internode spacing trait in soybeans; (2) Screening or breeding soybean single plants, lines, varieties or strains with long average internode spacing; (3) Screening or breeding soybean single plants, lines, varieties or strains with short average internode spacing; (4) Soybean breeding; (5) Prepare products for identification or auxiliary identification of the average internode spacing trait of soybeans; (6) Prepare or select soybean single plants, lines, strains or varieties with long average internode spacing; (7) Prepare or select soybean single plants, lines, varieties or strains with short average internode spacing; (8) Prepare soybean breeding products.

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