Wheat grain length molecular marker and application thereof

Abstract

The invention discloses a wheat grain length molecular marker and application thereof. The technical problem to be solved is how to provide a wheat grain length molecular marker. The invention specifically discloses any one of the following applications of an InDel molecular marker or a substance for detecting the InDel molecular marker: A1) an application in identification or auxiliary identification of wheat grain length; a2) application in preparation of products for identification or auxiliary identification of wheat grain length; a3) application in preparation of wheat breeding products; the InDel molecular marker is a DNA (deoxyribonucleic acid) molecule of which the nucleotide sequence is a sequence 5. The grain length of the deletion type wheat is longer than or longer than the grain length of the insertion type or hybrid type wheat, and the method can be used for industrial production.

Description

Technical Field

[0001] The invention relates to a wheat grain length molecular marker and application thereof in the field of biotechnology. Background Art

[0002] Wheat is one of the largest grain crops in the world, and its yield is directly related to world food security. Wheat grain yield is composed of the number of ears per unit area, the number of grains per ear, and grain weight. The coordination of the three is the key to achieving high wheat yields. Grain weight (1000-grain weight), as one of the yield components, is relatively stable and is relatively less affected by environmental conditions. It has always been valued by breeders, and improving grain weight is an important way to breed high-yield wheat.

[0003] Grain weight, as an important yield component, has relatively stable heredity and is composed of multiple subcomponents, including grain morphological parameters and grain filling rate. Grain morphological parameters can be decomposed into basic components such as grain length, grain width, and grain thickness. After wheat blooms and fertilizes, the fertilized egg first divides, and the endosperm nucleus is formed in the ovary and proliferates along the edge of the embryo sac. Finally, the embryo sac is filled with endosperm cells, and the proliferation of embryo and endosperm tissues is completed at about the same time. In terms of time, the increase in wheat grain volume is first in length, then in width and thickness. Generally speaking, 12-15 days after flowering, the grain length reaches the maximum value. After the length reaches the maximum value, the width and thickness increase rapidly, and at this time, the grain development is basically completed. After that, nutrients begin to accumulate in large quantities and enter the filling and maturity period. Wheat grain weight is affected by many factors during its formation process. In addition to genetic factors, it is also affected by external environmental factors such as light, temperature, and nutrients. Therefore, wheat grain yield is very susceptible to adverse external factors such as high temperature, drought and dry hot wind after flowering. High temperature, weak light and drought during the grain filling period will lead to a significant decrease in wheat thousand-grain weight. From a developmental perspective, the grain length is formed earlier than the width and thickness, so it is less affected by later environmental factors, and the trait expression is more stable, which is useful for breeding. Summary of the invention

[0004] The technical problem solved by the invention is how to identify or assist in identifying the length of wheat grains.

[0005] In order to solve the above technical problems, the present invention provides the following applications.

[0006] Any of the following applications of InDel molecule labeling or detecting a substance labeled with the InDel molecule:

[0007] A1) Application in identification or auxiliary identification of wheat grain length;

[0008] A2) Application in the preparation of products for identification or auxiliary identification of wheat grain length;

[0009] A3) Application in the preparation of wheat breeding products;

[0010] The InDel molecular marker is a DNA molecule having a nucleotide sequence of sequence 5.

[0011] In the present application, the indicator of grain length may be ten grains length or a single grain length.

[0012] The substance may be a product. The detection substance may include reagents, kits and instruments for detecting the above-mentioned InDel molecular markers. Specifically, primers and / or other reagents and instruments required for in vitro amplification of nucleic acids for detecting the above-mentioned InDel molecular markers.

[0013] In the present application, sequence 5 is the 592704585th to 622669697th position of the sequence with genebank number GenBank: CM031193.1 (06-MAY-2021).

[0014] In order to solve the above technical problems, the present invention also provides a product.

[0015] The product is a substance containing the above-mentioned InDel molecular marker, and can be any one of the following G1)-G3):

[0016] G1) detecting the product labeled by the InDel molecule;

[0017] G2) Application in identification or auxiliary identification of wheat grain length;

[0018] G3) Application in the preparation of products for identification or auxiliary identification of wheat grain length;

[0019] G4) in the preparation of wheat breeding products.

[0020] In order to solve the above technical problems, the present invention also provides a method for identifying or assisting in identifying the length of wheat grains.

[0021] The method comprises detecting wheat genotype to identify or assist in identifying wheat grain length:

[0022] The grain length of the wheat with the deletion type genotype is longer than the grain length of the wheat with the insertion type or heterozygous genotype. The deletion type is a homozygous type with the presence of the above-mentioned InDel molecular marker, the insertion type is a homozygous type without the above-mentioned InDel molecular marker, and the heterozygous type is a heterozygous type with the presence and absence of the above-mentioned InDel molecular marker.

[0023] In order to solve the above technical problems, the present invention also provides a method for wheat breeding.

[0024] The method comprises selecting homozygous wheat containing the above-mentioned InDel molecular marker in its genome as a parent for breeding.

[0025] In the above, the above method is applied in wheat breeding.

[0026] In the above-mentioned applications, products and methods, the purpose of the breeding includes cultivating or selecting wheat with long or short grain length.

[0027] In the above-mentioned application, product, method or application, product or method, the substance for detecting the InDel molecular marker is the following D1), D2), D3) or D4):

[0028] D1) in vitro nucleic acid amplification primers containing specific amplification InDel molecular markers;

[0029] D2) an in vitro nucleic acid amplification reagent containing the in vitro nucleic acid amplification primer described in D1);

[0030] D3) a kit containing the in vitro nucleic acid amplification primers described in D1) or the in vitro nucleic acid amplification reagents described in D2);

[0031] D4) A detection instrument containing the in vitro nucleic acid amplification primers described in D1), the in vitro nucleic acid amplification reagents described in D2) or the kit described in D3).

[0032] The in vitro nucleic acid amplification technology can be polymerase chain reaction (PCR), strand displacement amplification (SDA), ligase chain reaction (LCR) and nucleic acid sequence-dependent amplification (NASBA), rolling circle nucleic acid amplification (RCA), loop-mediated isothermal amplification (LAMP), helicase-dependent isothermal amplification technology (HDA) or Qβ replication technology.

[0033] The present application uses polymerase chain reaction (PCR) as an amplification method to perform polymorphism detection.

[0034] The specific amplification in D1) can detect the nucleotide sequence of the SNP polymorphic site by the presence or absence of an amplification product or by the presence or absence of an amplification product combined with an auxiliary reagent such as a probe.

[0035] In the above-mentioned application, product or method, the in vitro nucleic acid amplification primers include primer pair 1 and primer pair 2:

[0036] The primer pair 1 comprises a label 1-F and a label 1-R;

[0037] The primer pair 2 comprises a label 2-F and a label 2-R;

[0038] The nucleotide sequence of the marker 1-F is a single-stranded DNA of SEQ ID No. 2 in the sequence list;

[0039] The label 1-R is a single-stranded DNA whose nucleotide sequence is SEQ ID No. 3 in the sequence list;

[0040] The nucleotide sequence of marker 2-F is a single-stranded DNA of SEQ ID No. 4 in the sequence list;

[0041] The nucleotide sequence marked 2-R is a single-stranded DNA of SEQ ID No. 5 in the sequence list.

[0042] In order to solve the above technical problems, the present invention also provides a method for wheat breeding.

[0043] The method comprises performing deletion mutation on the sequence 5 region in the target wheat genome, and then obtaining deletion homozygous wheat to obtain wheat with a grain length longer than that of the target wheat.

[0044] In order to solve the above technical problems, the present invention also provides the following applications.

[0045] Application of the sequence of the wheat genome with genebank number GenBank: CM031193.1 (06-MAY-2021) or the sequence of positions 1 to 592704584 of the genebank number GenBank: CM031193.1 (06-MAY-2021) corresponding to different grain lengths in any of the following:

[0046] A1) Application in identification or auxiliary identification of wheat grain length;

[0047] A2) Application in the preparation of products for identification or auxiliary identification of wheat grain length;

[0048] A3) Application in preparing wheat breeding products.

[0049] In order to solve the above technical problems, the present invention also provides a DNA molecule.

[0050] The DNA molecule is a DNA molecule having a nucleotide sequence of position 592704585 to position 622669697 of sequence 5.

[0051] The in vitro nucleic acid amplification technology can be polymerase chain reaction (PCR), strand displacement amplification (SDA), ligase chain reaction (LCR) and nucleic acid sequence-dependent amplification (NASBA), rolling circle nucleic acid amplification (RCA), loop-mediated isothermal amplification (LAMP), helicase-dependent isothermal amplification technology (HDA) or Qβ replication technology.

[0052] The present application uses polymerase chain reaction (PCR) as an amplification method to perform polymorphism detection.

[0053] In the above-mentioned applications, methods and products, the in vitro nucleic acid amplification primer may be labeled or not labeled with a marker. The marker refers to any atom or molecule that can be used to provide a detectable effect and can be attached to a nucleic acid. Markers include but are not limited to dyes; radioactive labels, such as 32P; binding moieties, such as biotin; haptens, such as digoxin (DIG); luminescent, phosphorescent or fluorescent moieties; and fluorescent dyes alone or in combination with moieties that can inhibit or shift the emission spectrum by fluorescence resonance energy transfer (FRET). The marker can provide a signal that can be detected by fluorescence, radioactivity, colorimetry, weight determination, X-ray diffraction or absorption, magnetism, enzyme activity, etc. The marker can be a charged moiety (positive or negative charge) or, alternatively, can be charge neutral. The marker can include a nucleic acid or protein sequence or a combination thereof, as long as the sequence containing the marker is detectable. In some embodiments, the nucleic acid is directly detected without a marker.

[0054] The wheat may be at least one of the following:

[0055] The wheat may be a hybrid offspring of wheat A and wheat B, the InDel molecular marker is a naturally occurring deletion mutation, and both wheat A and wheat B are insertion-type. The hybrid offspring may be F2 generation and above, such as F2 generation, BC1F2, RIL population, etc.

[0056] The breeding objectives include developing wheat with longer kernel length.

[0057] The wheat A can be local yellow flower wheat. The wheat B can be Zhengmai 9023.

[0058] In the above applications, methods and products, the substance may be a reagent and / or kit and / or instrument required for determining the InDel molecular marker or genotype by at least one of the following methods: in vitro nucleic acid amplification, DNA sequencing, restriction fragment length polymorphism, single-stranded conformation polymorphism, denaturing high performance liquid chromatography and InDel chip. Among them, InDel chips include chips based on nucleic acid hybridization reaction, chips based on single base extension reaction, chips based on allele-specific primer extension reaction, chips based on "one-step" reaction, chips based on primer ligation reaction, chips based on restriction endonuclease reaction, chips based on protein DNA binding reaction, and chips based on fluorescent molecule DNA binding reaction.

[0059] Beneficial Effects

[0060] The present invention discloses a molecular marker for wheat grain length and its application, and solves the technical problem of how to provide a molecular marker for wheat grain length. Specifically disclosed is any one of the following applications of an InDel molecular marker or a substance for detecting the InDel molecular marker: A1) application in identifying or assisting in identifying wheat grain length; A2) application in preparing a product for identifying or assisting in identifying wheat grain length; A3) application in preparing a wheat breeding product; the InDel molecular marker is a DNA molecule with a nucleotide sequence of sequence 5. The grain length of the deletion type wheat is longer than that of the insertion type / heterozygous type wheat, and can be used for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0061] Figure 1 For materials GL, Beijing 411 and BC 3 F 4 Electrophoresis results of genotype detection of strains 1-8.

[0062] Figure 2 For materials GL, Beijing 411 and BC 3 F 4 Ten-grain length phenotype diagram of strains 1-8 genotype detection.

[0063] Figure 3 For materials GL, Beijing 411 and BC 3 F 4 Bar graph of grain length statistics for lines with insertion / heterozygous and deletion genotypes. DETAILED DESCRIPTION

[0064] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0065] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0066] The following examples were processed using SPSS11.5 statistical software, and the experimental results were expressed as mean ± standard deviation. One-way ANOVA test was used, and P < 0.05 (*) indicated a significant difference, P < 0.01 (**) indicated a very significant difference, and P < 0.001 (***) indicated an extremely significant difference.

[0067] The term "gene" refers to a segment of DNA involved in producing a polypeptide chain; it includes regions preceding and following the coding region (leader and trailer regions) that are involved in the transcription / translation of the gene product and the regulation of said transcription / translation, as well as intervening sequences (introns) between individual coding regions (exons).

[0068] The term "allele" refers to one of multiple alternative forms of a gene or non-coding region of DNA that occupies the same position on a chromosome. The term allele can be used to describe DNA from any organism, including but not limited to bacteria, viruses, fungi, protozoa, mold, yeast, plants, humans, non-humans, animals, and archaea.

[0069] The term "genotype" refers to the general term for all the genetic combinations of a biological individual. Biological individuals include, but are not limited to, diploids, tetraploids, or other possible polyploids. For example, in diploids, the genotype can be homozygous or heterozygous. A homozygous genotype means that the two alleles are exactly the same, such as AA or aa. A heterozygous genotype means that the two alleles are different, such as Aa. Other polyploid genotypes may appear and be described based on the actual situation of the alleles.

[0070] The term "InDel molecular marker" refers to an InDel marker (full name Insertion and Deletion marker), which is an insertion or deletion event that occurs in the genome, based on the insertion or deletion of nucleotides that occurs at a specific position in the genome. Similar to alleles, there can be multiple InDel molecular marker genotypes. For example, in diploids, the InDel molecular marker genotype can be homozygous or heterozygous. A homozygous genotype refers to two alleles that are exactly the same, such as AA or aa. A heterozygous genotype refers to two different alleles, such as Aa. Other polyploid genotypes may appear and be described based on the actual conditions of the alleles.

[0071] The term "template" refers to any nucleic acid molecule that can be used for amplification according to the present invention. Non-natural double-stranded RNA or DNA can be made into double-stranded DNA and used as double-stranded DNA. Any double-stranded DNA or product containing a variety of different double-stranded DNA molecules can be used as template DNA to amplify one or more loci contained in the template DNA.

[0072] The term "primer" refers to an oligonucleotide that can be used in an amplification method such as the polymerase chain reaction (PCR) for amplifying a nucleotide sequence based on a polynucleotide sequence that corresponds to a specific genomic sequence. At least one PCR primer used to amplify a polynucleotide sequence is sequence-specific for the sequence.

[0073] The term "amplification reaction" refers to a process for copying a nucleic acid one or more times. In embodiments, the amplification method includes, but is not limited to, polymerase chain reaction, self-sustained sequence reaction, ligase chain reaction, rapid amplification of cDNA ends, polymerase chain reaction and ligase chain reaction, Q-β phage amplification, strand displacement amplification, or overlap extension splicing polymerase chain reaction. In some embodiments, a single molecule nucleic acid is amplified, for example, by digital PCR.

[0074] The term "deletion mutation" refers to the process in which a codon encoding a certain amino acid becomes a codon encoding another amino acid after a base is deleted. The "deletion mutation" includes but is not limited to single base deletion, multiple base deletion, base fragment deletion, chromosome fragment deletion and chromosome deletion.

[0075] The term "homozygous deletion" refers to a chromosome fragment having a pair of deletion mutations in a chromosome structural variation of an individual, including but not limited to cells, cell lines, tissues, organs and complete individuals.

[0076] The term "wheat" refers to an annual or biennial herbaceous plant of the family Poaceae and the genus Triticeae.

[0077] The terms "include", "including", "have", "contain" and the like are all open terms, meaning including but not limited to.

[0078] The terms "first" and "second" are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of the features. Further, in the description of the present invention, unless otherwise specified, "plurality" means two or more.

[0079] When the term "identity" is used to describe an amino acid sequence or a nucleic acid sequence relative to a reference sequence, the percentage of identical amino acids or nucleotides between two amino acid sequences or nucleic acid sequences is determined by conventional methods, for example, see Ausubel et al., eds. (1995), Current Protocols in MolecμLar Biology, Chapter 19 (Greene Publishing and Wiley-Interscience, New York); and the ALIGN program (Dayhoff (1978), Atlas of Protein Sequence and Structure 5: Suppl. 3 (National Biomedical Research Foundation, Washington, DC). There are many algorithms for aligning sequences and determining sequence identity, including the homology alignment algorithm of Needleman et al. (1970) J. Mol. Biol. 48: 443; the local homology algorithm of Smith et al. (1981) Adv. Appl. Math. 2: 482; the local homology algorithm of Pearson et al. (1988) P. The similarity search method of Proc. Natl. Acad. Sci. 85: 2444; Smith-Waterman algorithm (Meth. Mol. Biol. 70: 173-187 (1997); and BLASTP, BLASTN, and BLASTX algorithms (see Altsch μL et al. (1990) J. Mol. Biol. 215: 403-410). Computer programs that utilize these algorithms are also available and include, but are not limited to, ALIGN or Megalign (DNASTAR) software, or WU-BLAS T-2 (Altsch μL et al., Meth. Enzym., 266:460-480 (1996)); or GAP, BESTFIT, BLAST Altsch μL et al., supra, FASTA, and TFASTA, available in the Genetics Computing Group (GCG) package, Version 8, Madison, Wisconsin, USA; and CLUSTAL in the PC / Gene program provided by Intelligenetics, Mountain View, California.

[0080] Material Description:

[0081] Zhengmai 9023: Xu Weigang. Zhengmai 9023, a new wheat variety approved by the state[J]. China Agricultural Information, 2009, (07): 34.

[0082] Local yellow wheat is recorded in the following document: "Chen Yaru, Zhang Qiaofeng, Fu Bisheng, Cai Shibin, Wu Jizhong, Chen Yahua. Analysis of differences in lead, cadmium and zinc accumulation in Chinese wheat micro-core germplasm grains and screening of low-accumulation varieties [J]. Journal of Nanjing Agricultural University, 2017, 40(03): 393-399.". The name in this document is local yellow wheat.

[0083] The deposit number of GL is CGMCC No.31802.

[0084] Biomaterial Name: GL

[0085] Variety name: Wheat

[0086] Latin name: Triticum aestivum

[0087] No.: CGMCC No.31802

[0088] Depository: China National Microbiological Culture Collection Administration General Microbiology Center

[0089] Abbreviation of depository institution: CGMCC

[0090] Address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing

[0091] Deposit date: October 12, 2024

[0092] CGMCC Registration Number: CGMCC No.31802

[0093] Example 1 Discovery of Molecular Markers

[0094] The molecular marker provided by the present invention is derived from a material GL (deposit number CGMCC No 31802) in the hybridization of local yellow flower wheat and wheat Zhengmai 9023 and continuous backcrossing with Zhengmai 9023. The grain length of GL is significantly increased. Through molecular marker identification and sequencing, it is found that the control of the grain length trait of the material is caused by the deletion of the terminal part of the long arm of its 6A chromosome.

[0095] As mentioned above, the present invention specifically discloses an Indel molecular marker, which is named GL, and the specific sequence is a 6A chromosome terminal deletion. The wheat 6A chromosome sequence is the sequence shown in the genebank number GenBank: CM031193.1 (06-MAY-2021). Sequence 5 is the 592704585-622669697 segment of the 6A chromosome sequence [genebank number GenBank: CM031193.1 (06-MAY-2021)].

[0096] The above molecular markers have three genotypes: deletion type, insertion type and heterozygous type.

[0097] Taking physical distance as the standard, the specific description is as follows:

[0098] The two homologous chromosomes in the A subgenome of the deletion type (also known as the deletion genotype) have the following characteristics: the DNA sequence from position 592704585 to position 622669697 (end) in genome 6A does not exist.

[0099] The two homologous chromosomes in the A subgenome of the insertion type (also called the insertion type genotype) have the following characteristics: the DNA sequence from position 592704585 to position 622669697 (end) in the genome 6A exists.

[0100] One homologous chromosome of the A subgenome of the heterozygous type (also called heterozygous genotype) has the following characteristics: the DNA sequence from position 592704585 to position 622669697 (end) in genome 6A is absent. The other homologous chromosome has the following characteristics: the DNA sequence from position 592704585 to position 622669697 (end) in genome 6A is present.

[0101] Taking the specific sequence as the standard, the specific description is as follows:

[0102] The two homologous chromosomes in the A subgenome of the deletion type (also called the deletion genotype) have the following characteristics: sequence 5 does not exist in the genome.

[0103] The two homologous chromosomes in the A subgenome of the insertion type (also known as the normal genotype) have the following characteristics: sequence 5 exists in the genome.

[0104] One homologous chromosome of the A subgenome of the heterozygous type (also called heterozygous genotype) has the following characteristics: sequence 5 does not exist in the genome. The other homologous chromosome has the following characteristics: sequence 5 exists in the genome.

[0105] Example 2 Development of Molecular Marker Primers

[0106] According to the above-mentioned deletion fragment sequence, the following molecular markers were designed as detection primers. The details of the molecular markers are shown in the following table (Table 1):

[0107] Table 1

[0108]

[0109] Genotyping methods:

[0110] The wheat genome was extracted and detected using marker 1-F / marker 1-R and marker 2-F / marker 2-R. The electrophoresis results showed that if the amplification result of marker 1-F / marker 1-R was about 697bp and the amplification result of marker 2-F / marker 2-R was about 561bp, the wheat genotype to be tested was an insertion type or a heterozygous type. If the amplification results of marker 1-F / marker 1-R and marker 2-F / marker 2-R were none, the wheat genotype to be tested was a deletion type. The amplified fragments were sequenced, and the results were the same as those of the above-mentioned genotype detection method. The above-mentioned established genotype detection method can be used for actual detection.

[0111] 1. Wheat Genomic DNA Extraction

[0112] The genomic DNA of wheat materials was extracted by conventional methods.

[0113] 2. PCR Amplification

[0114] 1) PCR reaction system (Novozyme, 2×Rapid Taq Plus Master Mix (Dye Plus))

[0115] Genomic DNA (100 ng / μL): 1 μL;

[0116] Primer F (10 μM): 1.0 μL;

[0117] Primer R (10 μM): 1.0 μL;

[0118] 2×PCR buffer for KOD FX: 12.5μL;

[0119] dH 2 O: 9.5 μL.

[0120] Example 3 Development and verification of molecular marker function

[0121] To verify the effect of the deletion on grain length, we hybridized the material (material GL, identified as deletion type by the above genotype detection method and genome sequencing) with the wheat variety Jing 411 (identified as insertion type by genome sequencing), and backcrossed with Jing 411 three times in a row, and then selfed four times to obtain BC 3 F 4 group.

[0122] The above BC 3 F 4 Populations, materials GL and Jing 411 were planted in the Changping Farm of the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, during the 2022-2023 growing season and grown under conventional field management conditions. When wheat grew to maturity, the grain length was measured (the results are shown in Tables 2 and Figure 3 ( Figure 3 In the above table, Jing 411 is the wheat variety Jing 411, GL is the material GL, BC 3 F 4 (Insertion or deletion) is BC 3 F 4 Wheat with insertion or deletion genotypes in the population, BC 3 F 4 (deletion type) is BC 3 F 4 The wheat genotype in the population is the missing type, the vertical axis unit is millimeter (mm), and the ten-grain length phenotype is recorded (the results are as follows Figure 2 As shown, Figure 2 In the above table, Jing 411 is the wheat variety Jing 411, GL is the material GL, BC 3 F 4 Strain 1, BC 3 F 4 Strain 2, BC 3 F 4 Strain 3, BC 3 F 4 Strain 4, BC 3 F 4 Strain 5, BC 3 F 4 Strain 6, BC 3 F 4 Strain 7 and BC 3 F 4 Strain 8 corresponds to BC 3 F 4 In the group Figure 1 BC 3 F 4 strains 1-8).

[0123] Grain length detection method:

[0124] After the wheat materials were harvested, the total length of 10 grains was randomly measured and converted into the length of a single grain after dividing by 10. Four replicates were performed, and 40 individual plants were measured for materials GL and Jing 411, and 40 individual plants for BC. 3 F 4 A total of 160 plant lines were measured, and the grain length analysis was performed using SPSS11.5 statistical software. The experimental results were expressed as mean ± standard deviation. One-way ANOVA test was used, and P < 0.05 (*) indicated a significant difference, P < 0.01 (**) indicated an extremely significant difference, and P < 0.001 (***) indicated an extremely significant difference.

[0125] Ten-grain length: After the wheat material was harvested, the total length of 10 grains was randomly measured in the grains, with 4 replicates. For materials GL and Jing 411, 40 individual plants were measured, and for BC 3 F 4 160 strains were measured. SPSS11.5 statistical software was used to process the data. The experimental results were expressed as mean ± standard deviation. One-way ANOVA test was used. P < 0.05 (*) indicated a significant difference, P < 0.01 (**) indicated an extremely significant difference, and P < 0.001 (***) indicated an extremely significant difference.

[0126] Among them, GL, Beijing 411 and BC 3 F 4 The ten-grain lengths of strains 1-8 are as follows: GL (8.6 cm), Jing 411 (7.5 cm), BC 3 F 4 Strain 1 (7.4 cm), BC 3 F 4 Strain 2 (7.7 cm), BC 3 F 4 Strain 3 (7.5 cm), BC 3 F 4 Strain 4 (7.3 cm), BC 3 F 4 Strain 5(8.6cm), BC 3 F 4 Strain 6 (8.5 cm), BC 3 F 4 Strain 7 (7.9 cm), BC 3 F 4 Strain 8 (7.4 cm).

[0127] The genotype detection method in Example 2 was used to perform genotype detection on the wheat planted above. The results are shown in Table 2 and Figure 1 ( Figure 1 In the above table, Jing 411 is the wheat variety Jing 411, GL is the material GL, BC 3 F 4 Strain 1, BC 3 F 4 Strain 2, BC 3 F 4 Strain 3, BC 3 F 4 Strain 4, BC 3 F 4 Strain 5, BC 3 F 4 Strain 6, BC 3 F 4 Strain 7 and BC 3 F4 Strain 8 corresponds to BC 3 F 4 In the group Figure 1 BC 3 F 4 strains 1-8) and found that BC 3 F 4 In the population offspring materials, the grain length of plants with the deletion marker (deletion type) was significantly longer than that of plants without deletion (insertion type or heterozygous type), indicating that the deletion fragment can increase grain length and can be applied to the genetic improvement of wheat grain type.

[0128] Table 2BC 3 F 4 Population genotype and grain length

[0129]

[0130]

[0131]

[0132]

[0133]

[0134] The present invention has been described in detail above. It will be apparent to those skilled in the art that the present invention may be implemented in a wide range under equivalent parameters, concentrations and conditions without departing from the spirit and scope of the present invention and without the need for unnecessary experimentation. Although the present invention provides specific embodiments, it should be understood that further improvements may be made to the present invention. In short, according to the principles of the present invention, this application intends to include any changes, uses or improvements to the present invention, including changes made by conventional techniques known in the art that depart from the scope disclosed in this application. Applications of some of the basic features may be made within the scope of the following appended claims.

Claims

1. Any of the following applications of InDel molecule labeling or detecting a substance labeled with the InDel molecule: A1) Application in identification or auxiliary identification of wheat grain length; A2) Application in the preparation of products for identification or auxiliary identification of wheat grain length; A3) Application in the preparation of wheat breeding products; The InDel molecular marker is a DNA molecule having a nucleotide sequence of sequence 5.

2. A product, characterized in that The product is a substance containing the InDel molecular marker for detecting the InDel according to claim 1, and can be any one of the following G1)-G3): G1) detecting the product marked by the InDel molecule; G2) Application in identification or auxiliary identification of wheat grain length; G3) Application in the preparation of products for identification or auxiliary identification of wheat grain length; G4) in the preparation of wheat breeding products.

3. A method for identifying or assisting in identifying wheat grain length, characterized in that: The method comprises detecting wheat genotype to identify or assist in identifying wheat grain length: The grain length of the wheat with the deletion type genotype is longer than or has the potential to be longer than the grain length of the wheat with the insertion type or heterozygous genotype. The deletion type is a homozygous type in which the InDel molecular marker described in claim 1 is present, the insertion type is a homozygous type in which the InDel molecular marker described in claim 1 is not present, and the heterozygous type is a heterozygous type in which the InDel molecular marker described in claim 1 is present and the InDel molecular marker described in claim 1 is not present.

4. A method for wheat breeding, comprising selecting a homozygous wheat containing the InDel molecular marker according to claim 3 in its genome as a parent for breeding.

5. The use according to claim 1, the product according to claim 2, the method according to claim 3 or 4, characterized in that: The breeding objectives include developing or selecting wheat with long or short grain length.

6. The use according to claim 1, the product according to claim 2, the method according to claim 4 or the use, product or method according to claim 5, characterized in that: The substance for detecting the InDel molecule marker is the following D1), D2), D3) or D4): D1) in vitro nucleic acid amplification primers containing specific amplification InDel molecular markers; D2) an in vitro nucleic acid amplification reagent containing the in vitro nucleic acid amplification primer described in D1); D3) a kit containing the in vitro nucleic acid amplification primers described in D1) or the in vitro nucleic acid amplification reagents described in D2); D4) A detection instrument containing the in vitro nucleic acid amplification primers described in D1), the in vitro nucleic acid amplification reagents described in D2) or the kit described in D3).

7. The use, product or method according to claim 6, characterized in that: The in vitro nucleic acid amplification primers include primer pair 1 and primer pair 2: The primer pair 1 comprises a label 1-F and a label 1-R; The primer pair 2 comprises a label 2-F and a label 2-R; The nucleotide sequence of the marker 1-F is a single-stranded DNA of SEQ ID No. 1 in the sequence list; The label 1-R is a single-stranded DNA whose nucleotide sequence is SEQ ID No. 2 in the sequence list; The nucleotide sequence of marker 2-F is a single-stranded DNA of SEQ ID No. 3 in the sequence list; The nucleotide sequence marked 2-R is a single-stranded DNA of SEQ ID No. 4 in the sequence list.

8. A method for wheat breeding, characterized in that: The method comprises performing deletion mutation on the sequence 5 region in the target wheat genome, and then obtaining deletion homozygous wheat to obtain wheat with a grain length longer than that of the target wheat.

9. Application of the sequence of the wheat genome with genebank number GenBank: CM031193.1 (06-MAY-2021) or the sequence from position 1 to position 592704584 of the genebank number GenBank: CM031193.1 (06-MAY-2021) corresponding to different grain lengths in any of the following: A1) Application in identification or auxiliary identification of wheat grain length; A2) Application in the preparation of products for identification or auxiliary identification of wheat grain length; A3) Application in preparing wheat breeding products.

10. A DNA molecule, characterized in that The DNA molecule is a DNA molecule having a nucleotide sequence of sequence 5.

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