Primer combination for water conservation and quality screening of winter wheat and application of primer combination

Abstract

The invention discloses a primer combination for water saving and quality screening of winter wheat and application thereof, and relates to the technical field of molecular biology, the primer combination for water saving and quality screening of winter wheat comprises a primer pair P1 and a primer pair P2; according to the primer combination, the water-saving and drought-resistant characters are screened through PCR amplification, and SDS-PAGE electrophoresis is combined to detect grain protein subunit composition so as to screen quality characters. The efficient breeding method suitable for winter wheat is formed by designing a specific primer combination, constructing a molecular marker screening system and innovatively combining a cultivation management mode of water before drought and drought after water before winter wheat. According to the method, dependence on a single known gene is avoided, oriented improvement of water-saving drought resistance and quality is realized through collaborative screening and environment adaptability cultivation of a primer combination, and a new technical path is provided for wheat breeding in the arid region of North China.

Description

Technical Field

[0001] This invention relates to the field of molecular biology, and in particular to a primer combination for water-saving and quality screening of winter wheat and its application. Background Technology

[0002] Hebei Province, as one of my country's major wheat-producing areas, has an annual wheat planting area of ​​over 33 million mu (approximately 2.8 million hectares), ranking among the top in the country in both yield per unit area and total output, and undertaking the important task of ensuring national food security. Its unique natural resource endowment has formed significant industrial advantages—abundant light and heat resources and fewer pests and diseases have nurtured high-quality medium-strong gluten wheat, making it a major high-quality wheat producing area in my country; "water conservation" and "high quality" have long been the core goals of wheat breeding in Hebei Province.

[0003] Traditional wheat breeding relies on field phenotypic screening, which suffers from problems such as long cycles, low efficiency, and difficulty in aggregating target traits. Although the development of molecular marker technology has provided means for detecting water-saving and drought-resistant genes and quality-related subunits, existing methods mostly focus on single gene functions and lack systematic solutions for the drought-stressed environment and high-quality wheat requirements of Hebei Province. For example, relying solely on marker screening based on known gene sequences makes it difficult to simultaneously consider field agronomic traits and regional adaptability; conventional cultivation management does not fully integrate the water regulation needs of arid areas, resulting in limited synergistic improvement effects of water conservation and high yield.

[0004] With the advancement of agricultural water-saving policies in Hebei Province, developing a breeding method that can efficiently combine water-saving and drought-resistant characteristics with high-quality grains has become an urgent technical problem to be solved. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a primer combination for water-saving and quality screening of winter wheat and its application.

[0006] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is as follows.

[0007] A primer set for water-saving and quality screening of winter wheat, the primer set including primer pair P1 and primer pair P2; the primer set screens water-saving and drought-resistant traits by PCR amplification and screens quality traits by SDS-PAGE electrophoresis to detect the composition of grain protein subunits.

[0008] As a preferred embodiment of the present invention, primer pair P1 is primer pair 1F and 1R composed of SEQ ID NO.1 and SEQ ID NO.2 in the sequence list; primer pair P2 is primer pair 2F and 2R composed of SEQ ID NO.3 and SEQ ID NO.4 in the sequence list.

[0009] A method for screening winter wheat germplasm resources based on the above primer combinations includes the following steps:

[0010] (1) Construct a winter wheat germplasm resource bank and collect locally approved or promoted wheat varieties;

[0011] (2) Extract genomic DNA from the germplasm resources, perform PCR amplification using the primer combination described in claim 1, and screen for strains that meet the following conditions:

[0012] The target fragment A was amplified using primer pair P1;

[0013] Fragment B, with a length of 677 bp, was amplified using primer pair P2;

[0014] (3) For the strains selected in step (2), the composition of seed protein subunits is further detected by SDS-PAGE electrophoresis to screen strains that meet the high-quality protein standard;

[0015] (4) Include strains that simultaneously meet the conditions of fragment A, fragment B and high-quality protein subunits into the molecular marker screening library.

[0016] A method for breeding a water-saving and high-quality winter wheat variety includes the following steps:

[0017] (1) Selecting parents from the molecular marker screening library described in claim 2 for hybridization;

[0018] (2) Extract single-plant DNA from the F2 generation population, perform PCR amplification using the primer combination of claim 1, and screen for homozygous single plants that simultaneously meet the following conditions:

[0019] The amplification product of fragment A was verified by sequencing to be consistent with the target region; the nucleotide sequence of the target region is shown in SEQ ID NO.5;

[0020] Fragment B is 677bp in length;

[0021] (3) The seed protein subunit composition of the single plants screened in step (2) was detected by SDS-PAGE electrophoresis, and the single plants with homozygous high-quality protein subunits were retained.

[0022] (4) The selected individual plants were cultivated in the field in the F3-F5 generation, and the management mode of watering before drought was adopted to obtain a stable strain with strong drought resistance and excellent quality.

[0023] Further preferred, the irrigation amount for the "water first, dry later" cultivation management in step (4) is 40-60m³. 3 / mu, no irrigation during the grouting and ripening stages.

[0024] A screening kit for water-saving and high-quality traits in winter wheat, comprising: the above primer combination, PCR reaction buffer, standard controls for identifying fragments A and B, and protein detection reagents for SDS-PAGE electrophoresis.

[0025] The beneficial effects of adopting the above technical solution are as follows: This invention, by designing specific primer combinations, constructing a molecular marker screening system, and innovatively combining it with the "pre-watering followed by drought" cultivation management model, forms a highly efficient breeding method suitable for winter wheat. This method avoids dependence on a single known gene, and instead achieves targeted improvement of water-saving and drought-resistant properties and quality through synergistic screening of primer combinations and environmental adaptability breeding, providing a new technical path for wheat breeding in the arid regions of North China. Attached Figure Description

[0026] Figure 1 This is a PCR amplification diagram of gene fragment A specificity, where M: DL2000 marker; 1-12 are different germplasm resources; 13: water;

[0027] Figure 2 Electrophoresis diagrams of the two alleles of fragment B;

[0028] Figure 3 Electrophoresis diagrams of high-quality protein subunits from some wheat varieties.

[0029] Figure 4 Sequence alignment and primer setup for amplified fragment A (expected length: 262 bp) were performed (a); the gene TraesCS4B02G072200.1 and its tandem repeat genes TraesCS4B02G072100.1 and TraesCS4B02G072300.1 were searched at https: / / www.wheatgenome.org / ; the search results and the seven gene sequences involved were compared using DNAStar software, and specific primers were designed using Primer Primer 5.0 at the differential sites (black boxes), which were synthesized by Shanghai Sangon Biotech. The upstream primer was 5'-CGGACCCGTGAAGTGCGCA-3'SEQ ID NO.1, and the downstream primer was WD40R1: 5'-GCGCTTCGGCTCCGAAATC-3'SEQ ID NO.2. Detailed Implementation

[0030] The following embodiments illustrate the present invention in detail. All raw materials and equipment used in the present invention are conventional commercially available products and can be directly obtained through market purchase. In the following description of the embodiments, specific details such as particular system structures and techniques are set forth for illustrative purposes and not for limitation, in order to provide a thorough understanding of the embodiments of this application. However, those skilled in the art will understand that the present application can also be implemented in other embodiments without these specific details. In other instances, detailed descriptions of well-known systems, apparatuses, and methods are omitted so as not to obscure the description of this application with unnecessary detail.

[0031] It should be understood that, when used in this specification and the appended claims, the term "comprising" indicates the presence of a described feature, integral, step, operation, element, and / or component, but does not exclude the presence or addition of one or more other features, integrals, steps, operations, elements, components, and / or a collection thereof. It should also be understood that the term "and / or" as used in this specification and the appended claims refers to any combination of one or more of the associated listed items and all possible combinations, and includes such combinations. As used in this specification and the appended claims, the term "if" may be interpreted, depending on the context, as "when," "once," "in response to determination," or "in response to detection." Similarly, the phrase "if determined" or "if detected [the described condition or event]" may be interpreted, depending on the context, as meaning "once determined," "in response to determination," "once detected [the described condition or event]," or "in response to detection [the described condition or event]."

[0032] 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. References to "one embodiment" or "some embodiments" in this application mean that one or more embodiments of this application include a specific feature, structure, or characteristic described in connection with that embodiment. Therefore, the phrases "in one embodiment," "in some embodiments," "in other embodiments," "in still other embodiments," etc., appearing in different parts of 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.

[0033] Example 1: Detection of target fragments in germplasm resources

[0034] The breeding methods for improving the water-saving and quality of winter wheat specifically include the following steps:

[0035] Primer design: Two pairs of specific primers were designed by analyzing target regions in the winter wheat genome that are associated with water conservation and drought resistance.

[0036] Primer pair P1 (F1: 5'-CGGACCCGTGAAGTGCGCA-3' (SEQ ID NO.1); R1: 5'-GCGCTTCGGCTCCGAAATC-3' (SEQ ID NO.2)): Targeted at the region on chromosome 4B, used to amplify fragment A (expected length: 262bp). Fragment A contains allelic variations, where the genotype of fragment A that is homozygous for CC is named C, and the genotype of fragment A that is homozygous for TT is named c;

[0037] Genomic DNA was extracted from 356 wheat germplasm resources as PCR templates. The PCR amplification system was 20 μL, containing 2 μL of DNA template (50 ng·μL⁻¹), 1 μL each of forward and reverse primers (10 μmol·L⁻¹), 10 μL of 2×Taq Master Mix (Sangon Biotech, Shanghai), and 6 μL of ddH₂O. The PCR reaction program was: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 40 s, 61.1℃ annealing for 40 s, 72℃ extension for 40 s, 35 cycles; 72℃ extension for 10 min. The PCR products were detected by agarose gel electrophoresis. Twelve PCR products were randomly selected for sequencing, and specific amplification of fragment A was successfully achieved. Figure 1 ).

[0038] The criteria for judging after sequencing are: the amplification product of fragment A is verified by sequencing to be consistent with the target region; the nucleotide sequence of the target region is shown in SEQ ID NO.5.

[0039] Primer pair P2 (F2: 5'-GGAGCATGCTTACAAAACCTATG-3' (SEQ ID NO.3); R2: 5'-GACCCACCTGTCAGAGACTTT-3' (SEQ ID NO.4)): specifically amplifies fragment B with a length of 677 bp targeting the target region of chromosome 2A;

[0040] Genomic DNA was extracted from 356 wheat germplasm resources as PCR templates. The PCR amplification system was 20 μL, containing 2 μL of DNA template (50 ng·μL⁻¹), 1 μL each of forward and reverse primers (10 μmol·L⁻¹), 10 μL of 2×Taq Master Mix (Sangon Biotech, Shanghai), and 6 μL of ddH₂O. The PCR reaction program was: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 40 s, 56.0℃ annealing for 40 s, 72℃ extension for 90 s, 35 cycles; 72℃ extension for 10 min. The PCR products were detected by agarose gel electrophoresis. Figure 2 The fragment sizes are 677bp (D) and 1378bp (d), respectively.

[0041] Verification by agarose gel electrophoresis and sequencing showed that all 12 samples randomly sequenced from fragment A showed ≥99% consistency.

[0042] In fragment B, the 677bp band accounts for 75.8% (270 / 356).

[0043] Based on the compositional characteristics of fragment A and fragment B, the 356 germplasm resources were divided into four haplotypes: CD, Cd, cD, and cd (Table 1).

[0044] Table 1. Types of Water-Saving and Drought-Resistant Genes

[0045]

[0046]

[0047]

[0048]

[0049]

[0050]

[0051]

[0052]

[0053]

[0054] According to Mendel's law of independent assortment, there are 16 types of offspring after the combination (Table 2). According to the breeding goal, the offspring materials are expected to contain two superior allelic variations, fragment A and fragment B. Among them, 9 parental combinations meet the requirements, namely CD×CD, CD×Cd, CD×cD, CD×cd, Cd×CD, Cd×cD, cD×CD, cD×Cd, and cd×CD.

[0055] Table 2 Parental Types of Water-Saving and Drought-Resistant Genes

[0056]

[0057] Example 2: Detection of high-quality protein subunits

[0058] (1) SDS-PAGE electrophoresis

[0059] Two proteins from each germplasm resource were individually ground into powder and subjected to SDS-PAGE electrophoresis. Based on the electrophoretic band positions of the control material Shi Luan 02-1, the high-quality protein bands of each germplasm resource were read to determine the subunit composition (e.g., Figure 3 );

[0060] (2) Composition of high-quality protein subunits in germplasm resources:

[0061] The results of high-quality protein testing of 356 germplasm resources showed that 217 germplasm resources contained the 7+8 high-quality subunit (Table 2). It is known that the proportion of the 7+8 high-quality subunit at the B1 locus in wheat from the Huang-Huai wheat region is over 60%, making it the dominant subunit. To improve breeding efficiency, high-quality subunits at the A1 and D1 loci were screened first based on the 217 materials containing the 7+8 subunit. A total of 11 types were detected, namely 1, 7+8, 5+10, 1, 7+8, 5+12, 1, 7+8, 2+12, 1, 7+8, 4+12, n, 7+8, 5+10, n, 7+8, 5+12, n, 7+8, 2+12, n, 7+8, 4+12, 2*, 7+8, 5+10, 2*, 7+8, 5+12, 2*, 7+8, 4+12 (Table 3).

[0062] The specific method for identifying high-quality protein subunits as parents is as follows: For A1 loci of 1, n, or 2*, subunit 1 is named E, and the rest are named e; for D1 loci of 5+10 and 5+12, they are named F, and the rest are named f. A1 and D1 loci are located on chromosomes 1A and 1D, respectively, conforming to the law of independent assortment, resulting in 16 offspring types (Table 4). Based on the breeding objective, the desired offspring materials contain either 1 and 5+10 or 1 and 5+12 superior allelic variations. Nine parental combinations meet this requirement: EF×EF, EF×Ef, EF×eF, EF×ef, Ef×EF, Ef×eF, eF×EF, eF×Ef, and ef×EF.

[0063] Table 3 Composition of high-quality protein subunits

[0064]

[0065]

[0066]

[0067]

[0068]

[0069] Table 4. Types of High-Quality Protein Parents

[0070]

[0071] Example 3, Configuration Combination

[0072] (1) Based on the principle that the two parents have complementary genes related to water conservation, drought resistance and quality, the combination should be configured. Among the germplasm resources, 171 materials contain excellent allelic variations of both fragment A and fragment B of water conservation and drought resistance genes. When using them as parents, only the high-quality subunit configuration combination determined in Example 2 needs to be referred to. Among the germplasm resources, 160 materials contain only one excellent allelic variation of water conservation and drought resistance genes. At the same time, the combination of water conservation and drought resistance genes and high-quality subunits in Example 1 should be referred to. Among the germplasm resources, 25 materials do not contain excellent allelic variations of water conservation and drought resistance genes. When using them as parents, they can only be combined with materials of type CD of water conservation and drought resistance genes. At the same time, the combination of high-quality subunits determined in Example 2 should be referred to.

[0073] (2) Water-saving, drought-resistant, and quality of wheat are complex agronomic traits controlled by multiple genes. Aggregating more water-saving, drought-resistant genes and quality-related loci is expected to improve the water-saving, drought-resistant, and quality of wheat. In order to breed water-saving, drought-resistant, and high-quality wheat varieties, it is also necessary to comprehensively consider field phenotypes, such as complementary combinations of agronomic traits such as high yield, yellowing tendency, disease resistance, and cold resistance. The combination of molecular design breeding and conventional selection is the basis for breeding new water-saving, drought-resistant, and high-quality varieties.

[0074] Example 4: Molecular Marker Tracking Detection

[0075] (1) The specific method for tracking and detecting water-saving and drought-resistant gene molecular markers is as follows: Select single plants with excellent agronomic traits from the F2 generation of each combination, mark them, extract gDNA from the leaves, and use the primers of Example 1 to perform PCR amplification on fragment A and fragment B respectively, and detect them by agarose gel electrophoresis. The number of F2 generation lines detected for each combination should not be less than 16.

[0076] (2) The specific method for tracking and detecting high-quality subunits is as follows: select single plants with excellent allelic variants of fragment A and fragment B in F2 generation and harvest seeds (CCDD). Randomly select 2 seeds from each line, grind them, extract seed protein and perform SDS-PAGE electrophoresis to identify lines with high-quality subunits of A1 and D1 sites (EEFF).

[0077] Example 5: Cultivation Management with Water Supply Before Dry Season

[0078] The specific method for the pre-irrigation followed by drought cultivation management is as follows: Selected homozygous F3-F5 generations are sown with sufficient soil moisture to ensure full and robust seedling formation, fully leveraging individual tillering potential. During the spring wheat jointing stage, topdressing is applied simultaneously with irrigation until maturity (generally, wheat fields are irrigated once more during the grain-filling stage). Later drought treatment facilitates the screening of drought-resistant progeny in arid environments. Phenotypic assessments of conventional agronomic traits such as field vigor, cold resistance, plant height, yellowing tendency, disease resistance, and grain plumpness are conducted during the growth period.

[0079] Example 6: Participating in national or regional trials of varieties in central and southern Hebei

[0080] The successful practices in participating in the regional and biological trials of wheat varieties in central and southern Hebei Province are as follows:

[0081] (1) Among the water-saving and drought-resistant germplasm resources, Jimai 22 is a wheat variety that has been widely promoted and applied, and has the characteristics of wide adaptability; Liangxing 99 has good yellowing and its appearance is loved by the people. Both have been tested and controlled in the northern part of the Yellow Sea; Shiyou 17 is a high-quality variety bred by Shijiazhuang Academy of Agricultural Sciences. This variety has a strong wheat aroma and is very suitable for making noodles and dumplings. Shiyou 17, Jimai 22 and Liangxing 99 are complementary in terms of agronomic traits and quality, which meets the conventional breeding parent selection conditions.

[0082] (2) The composition of water-saving and drought-resistant fragments and high-quality protein subunits in the varieties Shiyou 17, Jimai 22, and Liangxing 99 was detected. It was found that the genotypes of their water-saving and drought-resistant fragments were Cd, cD, and cD, respectively, and the genotypes of their high-quality subunits were EF, eF, and ef, respectively, all of which showed complementarity. The genotype of Shiyou 17 is CdEF, the genotype of Jimai 22 is cDeF, and the genotype of Liangxing 99 is cDef (Table 5). The combinations of Shiyou 17 with Jimai 22 and Liangxing 99 were CdEF×cDeF and CdEF×cDef, respectively. The probability of CDEF being produced in the F2 generation of the CdEF×cDeF combination is 1 / 8, and the probability of CDEF being produced in the F2 generation of the CdEF×cDef combination is 1 / 64. Therefore, the Shiyou 17 / Jimai 22 combination should screen for no less than 8 F2 single plants, and the Liangxing 99 / Shiyou 17 combination should screen for no less than 64 F2 single plants. Marking and testing should continue until the F2 homozygous CDEF line is found and the single plant shows excellent performance in the field.

[0083] Table 5. Breeding varieties and their parental genotypes

[0084]

[0085] (3) F3-F5 strains with excellent agronomic traits were further screened under the pre-irrigation and post-dry cultivation model to participate in regional trials organized in the northern Huang-Huai region or the central and southern Hebei region. Shi 4366 and Shimai 26 successfully completed the trial process and obtained variety approval certificates.

[0086] Example 7, Quality Inspection

[0087] Protein characteristics are closely related to the quality of steamed buns, affecting their appearance, volume, specific volume, and internal structure. Specific quality testing methods include:

[0088] (1) The protein content of flour was determined by the Kjeldahl method for the approved varieties Shi 4366 and Shimai 26, the high-quality parent donor Shiyou 17, and the water-saving control variety Shimai 22. The relative content of proteins of different molecular weights in the flour, including SDS-insoluble protein macromers (UPP), high molecular weight polymeric proteins (LPP), high molecular weight monomeric proteins (LMP), low molecular weight monomeric proteins (SMP), and LPP / LMP ratio, was determined by an Agilent 1200 high performance liquid chromatograph (Agilent Technologies). Data were obtained from three replicates. The data were processed and plotted using Excel 2016, and the significance of differences was analyzed using SPSS 18.0 (ANOVA).

[0089] (2) Statistical analysis revealed that among the six protein content indicators, the PC and LMP content of Shimai 26 was significantly higher than that of Shiyou 17, and the PC, UPP, LPP and LPP / LMP of Shi4366 were significantly higher than those of Shiyou 17 (Table 6). The analysis of the texture parameters of steamed buns showed that the hardness, elasticity and resilience of Shimai 26 were significantly better than those of Shiyou 17. Except for resilience, Shi4366 was not significantly different from Shiyou 17, and its resilience was significantly better than that of the water-saving control variety Shimai 22 (Table 7).

[0090] Table 6. Relative content of proteins with different molecular weights

[0091]

[0092] Notes: Protein content (PC), SDS-insoluble protein macromers (UPP), high molecular weight polymeric protein (LPP), high molecular weight monomeric protein (LMP), low molecular weight monomeric protein (SMP), and LPP / LMP.

[0093] Table 7 Texture parameters of steamed buns made from wheat varieties

[0094]

[0095] This invention designs specific primer combinations to construct a molecular marker screening system and innovatively combines it with a "water-first, drought-second" cultivation management model, forming a highly efficient breeding method suitable for winter wheat. This method avoids dependence on a single known gene, instead achieving targeted improvement of water-saving and drought-resistant properties and quality through synergistic screening of primer combinations and environmental adaptability breeding, providing a new technical path for wheat breeding in the arid North China region.

[0096] 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.

[0097] 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. A primer combination for water saving and quality screening of winter wheat, characterized in that, The primer combination comprises primer pair P1 and primer pair P2; the primer combination is used for screening water-saving drought resistance traits by PCR amplification, and screening quality traits by detecting grain protein subunit composition by SDS-PAGE electrophoresis.

2. The primer combination for water-saving and quality screening of winter wheat according to claim 1, characterized in that: The primer pair P1 is primer pair 1F and 1R consisting of SEQ ID NO. 1 and SEQ ID NO. 2 in the sequence listing; and the primer pair P2 is primer pair 2F and 2R consisting of SEQ ID NO. 3 and SEQ ID NO. 4 in the sequence listing.

3. A method for screening winter wheat germplasm resources based on the primer combination of claim 1, characterized in that, The method comprises the following steps: (1) constructing a winter wheat germplasm resource library, collecting local approved or popularized wheat varieties; (2) extracting genomic DNA of the germplasm resources, performing PCR amplification using the primer combination of claim 1, and screening lines meeting the following conditions: amplifying target fragment A by primer pair P1; amplifying fragment B with a length of 677 bp by primer pair P2; (3) further detecting grain protein subunit composition of the lines screened in step (2) by SDS-PAGE electrophoresis, and screening lines meeting the high-quality protein standard; (4) including lines meeting the conditions of fragment A, fragment B and high-quality protein subunit in the molecular marker screening library.

4. A method for breeding a water-saving high-quality line of winter wheat, characterized in that, The method comprises the following steps: (1) selecting parents for hybridization from the molecular marker screening library of claim 2; (2) extracting single plant DNA in the F2 generation population, performing PCR amplification using the primer combination of claim 1, and screening homozygous single plants meeting the following conditions simultaneously: the amplification product of fragment A is verified by sequencing to be consistent with the target region; the length of fragment B is 677 bp; (3) retaining single plants homozygous for high-quality protein subunits by detecting grain protein subunit composition of the single plants screened in step (2) by SDS-PAGE electrophoresis; (4) cultivating the screened single plants in F3-F5 generations in the field, using the early water and late drought management mode, to obtain stable lines with strong drought resistance and excellent quality.

5. The selection method according to claim 4, wherein The irrigation amount of the "early water and late drought" cultivation management in step (4) is 40-60 m³ / acre, and irrigation is not performed during the grain filling period and the maturation period.

6. A kit for screening water saving quality traits in winter wheat, characterized in that, It comprises the primer combination of claim 1, PCR reaction buffer, standard controls for identifying fragments A and B, and protein detection reagents required for SDS-PAGE electrophoresis.

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