Application of soybean synergistic function module and target gene in improvement of salt tolerance of leguminous plants
By constructing the soybean miRNA-target gene functional module gma-miR482e/Glyma.12G236500 and regulating the expression of Glyma.12G236500, the problem of insufficient salt tolerance in soybeans was solved, and the yield and quality of soybeans in saline-alkali soil environments were improved.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-29
- Publication Date
- 2026-04-07
AI Technical Summary
Soybean growth is affected by soil salinization, resulting in yield loss. Current technologies lack effective molecular breeding methods to improve its salt tolerance.
By constructing soybean miRNA-target gene functional modules gma-miR482e and Glyma.12G236500, the expression level of Glyma.12G236500 was regulated to improve the salt tolerance of soybean. The methods included overexpressing Glyma.12G236500 or inhibiting the activity of gma-miR482e.
It significantly improves the yield and quality of soybeans in saline-alkali soil environments, provides new targets and technical support for molecular breeding, and has important application value.
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Figure CN121801943A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the field of plant biotechnology, and particularly relates to a soybean miRNA-target gene function module and application of a target gene in improving salt tolerance of leguminous plants, in particular to a regulatory relationship between gma-miR482e and Glyma.12G236500 and application value of the same in salt-tolerant breeding. BACKGROUND
[0002] Soybean is a major food crop in China, providing an important source of vegetable oil for residents' diet and a key supply of feed protein, playing a vital role in agricultural production. In 2024, the national soybean planting area was about 150 million mu, of which 1.233 million mu was planted in Xinjiang, with a total output of 20.65 million tons and an average yield of 133.3 kg / mu. However, data shows that China's soybean consumption still relies on imports by about 80%-85%, with an import volume of about 105 million tons in 2024.
[0003] As a seasonal crop, soybean growth depends on stable climate and soil conditions. In recent years, global climate anomalies have intensified, with frequent extreme high temperatures, droughts, and soil salinization, seriously disrupting the soybean growth cycle and restricting its yield and quality improvement. Among them, soil salinization has become a major environmental stress factor facing global agriculture, affecting about 20% of cultivated land. Salinization destroys the plant's living environment, leading to crop yield reduction or even complete loss. Soybean, as a salt-sensitive crop, is easily affected by salt damage during the entire growth period, resulting in significant yield loss. Therefore, to alleviate the impact of salinization on soybean production and expand planting area and improve yield are the main ways to increase production, and in-depth understanding of the causes of salinization, clarifying its harmful mechanism to plants, and systematically revealing the physiological and molecular basis of plant salt tolerance response have become the key to addressing this challenge.
[0004] The application integrates multi-omics data such as degradation group, transcriptome, and small RNA sequencing to discover the key regulatory module gma-miR482e-Glyma.12G236500 in salt stress response, and elucidates that gma-miR482e negatively regulates the expression of Glyma.12G236500, thereby affecting soybean salt tolerance. This achievement provides new evidence for analyzing the molecular network of plant salt stress response and provides new targets and technical support for genetic improvement and modular breeding of legume crops. SUMMARY
[0005] One of the purposes of the application is to provide a salt tolerance regulatory module composed of soybean microRNA gma-miR482e and its target gene Glyma.12G236500.
[0006] The second object of the present application is to provide the application of the regulatory module in regulating the salt tolerance of legume plants.
[0007] The third object of the present application is to provide a method for improving the salt tolerance of legume plants based on the target gene Glyma.12G236500.
[0008] To achieve the above-mentioned objects, the technical solutions of the present application are as follows:
[0009] A soybean synergistic functional module, which comprises a soybean microRNA gma-miR482e and its target gene Glyma.12G236500. Based on the degradation group, transcriptome and small RNA sequencing analysis, it is confirmed that gma-miR482e can specifically target and negatively regulate the expression of the target gene Glyma.12G236500; wherein the mature sequence of gma-miR482e is shown as SEQ ID NO. 1, and the coding sequence of Glyma.12G236500 is shown as SEQ ID NO. 2.
[0010] The sequence of the above-mentioned target gene Glyma.12G236500 also includes a gene derived therefrom formed by substitution, deletion or addition of one or more (for example, 1-30; preferably 1-20; more preferably 1-10, such as 5, 3) nucleotide residues, and has the same or similar salt tolerance regulation function as Glyma.12G236500; or a gene derived therefrom having 80% (preferably more than 90%, such as 95%, 98%, 99% or more) homology with the nucleotide sequence defined by Glyma.12G236500 and having the salt tolerance function of Glyma.12G236500.
[0011] The most important object of the present application is to provide the above-mentioned regulatory module with the function of regulating the salt tolerance of plants. It mainly relates to the application of enhancing the salt tolerance of legume plants by regulating the expression level of gma-miR482e or Glyma.12G236500 in the module.
[0012] In the process of identifying its function, the present application successfully obtained transgenic materials overexpressing Glyma.12G236500 through Agrobacterium-mediated soybean hairy root genetic transformation system. The results of salt tolerance phenotype analysis showed that overexpression of the target gene Glyma.12G236500 could significantly improve the salt tolerance of soybean. It is indicated that in specific practical application, the salt tolerance of legume plants can be improved by increasing the expression level of Glyma.12G236500, or inhibiting the activity of its negative regulatory factor gma-miR482e, thereby improving the yield and quality of soybean in saline-alkali environment.
[0013] Preferably, the legume is soybean. Specifically, the application is to improve the salt tolerance of soybean.
[0014] In addition, the present application also provides a method for improving the salt tolerance of plants, which can be method (1) or method (2):
[0015] Method (1) is to obtain a plant with improved salt tolerance by increasing the expression level and / or activity of the target gene Glyma.12G236500 in the plant of interest.
[0016] Method (2) is to obtain a plant with improved salt tolerance by reducing the expression level and / or activity of gma-miR482e in the plant of interest, thereby relieving its inhibition on Glyma.12G236500.
[0017] In the present application, the plant suitable for the present application or the plant of interest is not particularly limited, as long as it is suitable for genetic transformation operation, such as various crops, ornamental plants, or forestry plants, etc. The plant can be, but is not limited to, dicotyledon, monocotyledon or gymnosperm.
[0018] As a preferred way, the "plant" includes but is not limited to legume, especially soybean, and any plant with the gene or homologous gene is suitable.
[0019] Preferably, the plant of interest is soybean.
[0020] Preferably, the method (1) for increasing the expression of Glyma.12G236500 in the plant of interest is to introduce an expression vector containing the coding sequence of Glyma.12G236500 into the plant of interest.
[0021] Preferably, the method (2) for reducing the activity of gma-miR482e in the plant of interest is selected from the following: introducing an antagonist of gma-miR482e, constructing a silencing vector, CRISPR / Cas9-mediated gene editing or methylation modification, wherein one of the implementation ways is to introduce a nucleic acid molecule (such as short tandem target mimic STTM) for silencing or inhibiting gma-miR482e into the plant of interest.
[0022] Further, the way of introducing Glyma.12G236500 into the plant of interest includes the following steps:
[0023] Step one, construction of Glyma.12G236500 overexpression vector; including soybean root RNA extraction and reverse transcription, using the synthesized cDNA as template to carry out PCR amplification to obtain Glyma.12G236500 cDNA full-length sequence, and constructing Glyma.12G236500 overexpression vector; step two, soybean hairy root genetic transformation.
[0024] Preferably, in step one, the overexpression vector pZP211 is used to prepare Glyma.12G236500 overexpression vector pZP211-35S-Glyma.12G236500, and the pZP211-35S-Glyma.12G236500 plasmid is transferred into Agrobacterium rhizogenes to carry out soybean hairy root genetic transformation.
[0025] Preferably, in step two, the Agrobacterium containing the Glyma.12G236500 overexpression plasmid is used to infect the soybean cotyledon node, and after the hairy roots grow, the cotyledon node infection site and the part below it are buried with vermiculite, and water is poured to soak, and after culture, the plant overexpressing Glyma.12G236500 is obtained.
[0026] The "plant" mentioned in the present application includes whole plants, parent and progeny plants and different parts of plants, including seeds, fruits, buds, stems, leaves, roots (including tubers), flowers, tissues and organs, and the gene or nucleic acid of interest is present in these different parts. The "plant" mentioned herein also includes plant cells, suspension cultures, callus tissue, embryos, meristematic regions, gametophytes, sporophytes, pollen and microspores, and each of the foregoing objects also contains the gene / nucleic acid of interest.
[0027] The present application includes any plant cell, or any plant obtained or obtainable by the methods therein, and all plant parts and propagules thereof. The patent also includes transfected cells, tissues, organs or whole plants obtained by any of the foregoing methods. The only requirement is that the progeny exhibit the same genotypic or phenotypic characteristics as obtained using the methods in the present patent.
[0028] The present application also extends to harvestable parts of the plants as described above, but is not limited to seeds, leaves, fruits, flowers, stems, roots, rhizomes, tubers and bulbs. It also further relates to other derivatives of the plants after harvesting, such as dried granules or powders, oils, fats and fatty acids, starches or proteins. The present application also relates to foodstuffs or food additives obtained from the relevant plants.
[0029] The beneficial effects of the present application are that:
[0030] (1) The application finds the miRNA gma-miR482e differentially expressed in the salt treatment of Zihuang 34 and Dongnong 50 varieties through analysis, and also finds the specific target gene Gma.12G236500 of gma-miR482e, and gma-miR482e can specifically target and negatively regulate the expression of Gma.12G236500.
[0031] (2) The application first proposes that a soybean miRNA-target gene functional module can regulate soybean salt tolerance, in which gma-miR482 can negatively regulate / inhibit the expression of the target gene Gma.12G236500. It is proved that overexpression of the target gene Gma.12G236500 of the soybean miRNA-target gene functional module can significantly improve the salt tolerance of soybean, so it can be applied to soybean production and molecular breeding in the future through transgenic, molecular markers and other means, and has very important application value.
[0032] (3) The breeding method provided by the application can significantly improve the salt tolerance of soybean, has important theoretical value and practical significance for breeding high-quality and high-yield soybean germplasm, and has a broad application prospect in plant molecular breeding. BRIEF DESCRIPTION OF DRAWINGS
[0033] The drawings accompanying the specification of the application are used to provide further understanding of the application, the illustrative embodiments of the application and the description thereof are used to explain the application, and do not constitute undue limitation on the application.
[0034] Figure 1 T-plot graph in degradome sequencing.
[0035] Figure 2 Expression pattern of Gma.12G236500 in overexpressing soybean hairy roots.
[0036] Figure 3 Comparison of relative root elongation rates of soybean chimeric plants of empty vector (EV) and Glyma.12G236500 overexpressing strain; wherein a is the water treatment condition (0mM NaCl); b is the 120mM NaCl treatment condition, and the total number of repetitions is three.
[0037] Figure 4 Comparison of survival rates of soybean chimeric plants of empty vector (EV) and Glyma.12G236500 overexpressing strain.
[0038] Figure 5Comparison of the growth status of empty vector (EV) and Glyma.12G236500 overexpression soybean chimera plants; wherein a is the water treatment condition (0 mM NaCl); b is the 120 mM NaCl treatment condition. DETAILED DESCRIPTION
[0039] It should be noted that the following detailed description is exemplary in nature and is intended to provide further description of the application. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.
[0040] Unless otherwise indicated, the techniques employed in the examples are standard techniques well known to those of ordinary skill in the art. The experimental methods in the following examples are conventional unless otherwise indicated. Unless otherwise specified, the reagents and materials used are commercially available.
[0041] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Also, any method and material similar or equivalent to those described herein can be used in the practice of the present application. The described preferred methods and materials are presented as examples only.
[0042] Unless otherwise indicated, the practice of the present application will employ, unless otherwise indicated, plant biological, microbiological, molecular biological, biochemical, chemical, and biotechnological techniques, which are well known to persons of ordinary skill in the art. Such techniques are explained fully in the literature. In addition, the methods of DNA extraction, phylogenetic tree construction, gene editing methods, construction of gene editing vectors, and obtaining of gene edited plants, other than those employed in the following examples, are well known in the art and are described in the literature.
[0043] As used herein, the term "gene" can include introns and exons in genomic sequences, and / or can include coding sequences in cDNA, and / or can include cDNA and its regulatory sequences. In particular embodiments, such as with respect to isolated nucleic acid sequences, it is preferred that the cDNA be assumed.
[0044] "Expression vector" refers to a vector that has been modified to include expression elements (e.g., promoters, RBS, terminators, etc.) that allow the expression of a gene of interest.
[0045] "Agrobacterium-mediated transformation" refers to a technique in which a gene of interest is inserted into a modified T-DNA region, and the exogenous gene is transferred and integrated into plant cells by infection with Agrobacterium, and then transgenic plants are regenerated by cell and tissue culture techniques.
[0046] Obtaining of the fragment of the target gene Glyma.12G236500 of the miRNA-target gene functional module In the previous study, based on the root tissues of the salt-tolerant soybean variety Qihuang 34 (QH34) and the salt-sensitive soybean variety Dongnong 50 (DN50) under salt stress treatment (0, 2, 4, 8 hours), by integrating small RNA sequencing, degradation group sequencing and transcriptome data analysis, a soybean root miRNA-target gene interaction network under salt stress was systematically constructed. Through modular analysis and gene differential expression screening of the network, multiple core regulatory modules responding to salt stress were found. Further analysis showed that one of the regulatory modules composed of gma-miR482e and its target gene Glyma.12G236500 showed the most significant expression difference and regulatory specificity between the salt-tolerant and sensitive varieties. The degradation group sequencing data clearly verified the specific cleavage site of gma-miR482e on the Glyma.12G236500 transcript, confirming the direct target regulation relationship between them (as shown in Figure 1 ). Gma-miR482 can negatively regulate / inhibit the expression of the target gene Glyma.12G236500, based on which we named the functional module as gma-miR482e / Glyma.12G236500 salt-tolerance regulatory module.
[0047] The fragments of gma-miR482e and Glyma.12G236500 were obtained by bioinformatics methods, the sequence of gma-miR482e is shown as SEQ ID NO. 1, and the sequence of Glyma.12G236500 is shown as SEQ ID NO. 2. The full-length coding frame nucleotide sequence of the gene is 3084 bp in length;
[0048] SEQ ID NO. 1:
[0049]
[0050] SEQ ID NO. 2:
[0051]
[0052]
[0053]
[0054] Example 2 Soybean transgenic plants overexpressing Glyma.12G236500 have enhanced salt tolerance
[0055] I. Construction of Glyma.12G236500 overexpression vector
[0056] (1) Extraction and reverse transcription of soybean root RNA
[0057] 1. Total RNA extraction by TRIzol method
[0058] (1) Take about 0.1 g of soybean W82 root system, freeze in liquid nitrogen and grind into fine powder, add 1 mL of TRIzol extraction solution, vortex for 2 min, and stand at room temperature for 5 min;
[0059] (2) Centrifuge at 12000 rpm for 10 min at 4°C, remove the precipitate;
[0060] (3) Transfer the supernatant to a new centrifuge tube, add 200 μL of chloroform, vortex, and stand at room temperature for 3 min;
[0061] (4) Centrifuge at 11000 rpm for 10 min at 4°C;
[0062] (5) Transfer the supernatant to a new centrifuge tube, add 600 μL of isopropanol, and stand at room temperature for 10 min;
[0063] (6) Discard the supernatant, add 1 mL of 75% ethanol, resuspend, centrifuge at 11000 rpm for 5 min at 4°C; repeat once.
[0064] (7) Dry at room temperature for 5-7 min, dissolve the precipitate with 20 μL of DEPC-H2O, and store at -80°C.
[0065] 2. Reverse transcription of cDNA
[0066] (1) Use the kit provided by Vazyme Company to perform reverse transcription according to the instructions, take the RNA product of the previous step as the template, and perform the reaction in a 0.2 ml centrifuge tube, a total of two steps. The genomic gDNA removal mixed system is shown in Table 1, first prepare the genomic gDNA removal mixed system (Table 1), add 4xgDNA wiper Mix 4 μL, RNA template 1 pg-1 μg, RNase free ddH2O to 16 μL, mix gently with a pipette gun, centrifuge, and react on a PCR instrument at 42°C for 2 min:
[0067] Table 1 Genomic gDNA removal mixed system
[0068] 4 x gDNA wiper Mix 4 μL RNA 1 g g RNase free ddH2O qsp to 16 μL
[0069] (2) Take the reaction product of the previous step and add 5xHiScript III qRT SuperMix 4 μL to 20 μL, mix gently with a gun head, place the microcentrifuge tube on a PCR instrument at 37°C for 15 min; n; 85°C, 5s; the product is cDNA, which is taken out after the reaction and stored at -20°C.
[0070] (Three) Construction of Glyma.12G236500 overexpression vector
[0071] Design specific primer Glyma.12G236500 OX -F, Glyma.12G236500 oX -R, F, R end primers with BamHI restriction sites respectively. The reverse transcription synthesized cDNA as template for PCR amplification.
[0072] Glyma.12G236500 o x -F:
[0073] 5'-GGTACCCGAGGATCCATGGGTCGACCTAGA-3';
[0074] Glyma.12G236500 oX -R:
[0075] 5'-GTAGTCCATTCTAGAATCATACACGACGAT-3';
[0076] Wherein, the PCR amplification system is shown in Table 2.
[0077] PCR reaction program: 95℃ pre-denaturation 7min; 95℃ denaturation 30s, 56℃ annealing 1min, 72℃ extension 1min, 35 cycles; 72℃ extension 5min.
[0078] Table 2 PCR amplification system
[0079]
[0080]
[0081] The PCR product is electrophoresed by 1.5% agarose gel to obtain the cDNA full-length sequence of Glyma.12G236500 with the length of 3081bp.
[0082] (Three) Construction of Glyma.12G236500 overexpression vector
[0083] The target band was recovered using Kangwei DNA recovery and purification kit, and then the purified DNA fragment was connected to vector pGEM-T (Promega Corporation), and E. coli DH5a competent cells were transformed, positive clones were selected for plasmid extraction, and sequencing was completed by Shenguo Bioengineering (Shanghai) Co., Ltd. A Glyma.12G236500 fragment with a length of 308 Ibp was obtained, and the DNA sequence had a sequence table SEQ ID NO. 2. The purified cDNA fragment was connected to the overexpression vector pZP211 by single enzyme digestion (BamHI), and E. coli DH5a competent cells were transformed by heat shock, and colony PCR was performed on the recombinants, and agarose gel electrophoresis was detected. The recombinants contained a band with the same size as the target fragment. After the identified recombinants were cultured and the plasmid was extracted, the plasmid sequencing was compared with the original sequence, the connected fragment was the full-length cDNA and had no base mutation and deletion, proving that the Glyma.12G236500 overexpression vector pZP211-35S-Glyma.12G236500 was successfully constructed. The pZP211-35S-Glyma.12G236500 plasmid and the pZP211 empty vector plasmid were respectively transferred into Agrobacterium rhizogenes K599 for soybean hairy root genetic transformation.
[0084] II. Soybean hairy root genetic transformation
[0085] (1) Soybean Williams 82 was planted in vermiculite and germinated in a greenhouse. When the 6-day-old cotyledon was not fully unfolded, the cotyledon node was infected with Agrobacterium containing Glyma.12G236500 overexpression plasmid and Agrobacterium containing pZP211 empty vector plasmid, respectively.
[0086] (2) After injection, cover with a transparent cover and keep the inside humid. After the hairy roots grow, bury the soybean cotyledon node and the part below it with vermiculite, and water thoroughly.
[0087] (3) Remove the transparent cover after 6 days, cover the infected part and the part below it with wet vermiculite to maintain a humid environment, and water every 2 days. 28°C, 14h light / 10h dark, cultivate for 3 weeks or so for salt tolerance phenotype analysis.
[0088] III. Positive identification of overexpression transgenic materials
[0089] Take Glyma.12G236500 overexpression plant single root, extract genomic DNA, using the specific primers of Glyma.12G236500 used to construct the expression vector to amplify, and the PCR product is identified by agarose gel electrophoresis. If a clear band is obtained, it is a Glyma.12G236500 overexpression transgenic positive seedlings, which can be used for phenotype analysis.
[0090] Four, Glyma.12G236500 expression pattern detection in soybean hairy roots overexpression
[0091] The expression pattern of Glyma.12G236500 in soybean hairy roots overexpression was detected by qRT-PCR method. The transgenic positive seedlings of Glyma.12G236500 and empty vector were taken from the root system, and then frozen in liquid nitrogen and stored in-80℃ refrigerator. The total RNA was extracted by TRIzol method and the cDNA was obtained by reverse transcription. The reaction was carried out by using the fluorescence quantitative kit of TAKARA company. The reaction was carried out on the quantitative PCR instrument (Applied Biosystems Stepone Plus), and the expression of the gene was detected according to the relative quantitative method. The reaction program was carried out according to the operation manual provided by TAKARA, and the soybean Actin gene was used as the internal reference in the reaction. The primer sequence is:
[0092] GmActin-F: 5'-CGGTGGTTCTATCTTGGCATC-3'
[0093] GmActin-R: 5'-GTCTTTCGCTTCAATAACCCTA-3'
[0094] Glyma.12G236500-F: 5'-GAACGGGAAGCAGAAGTGGA-3'
[0095] Glyma.12G236500-R: 5'-GGGACAATCATTTCGCCAGC-3'
[0096] The reaction program is as follows: pre-denaturation 94℃ 5min, denaturation 94℃ 30s, annealing 56℃ 40s, extension 72℃ 40s, 35 cycles of reaction. After the end, 2 -ΔΔCt Method to calculate the expression of Glyma.12G236500. This experiment was repeated three times. The results are shown in Figure 2 Glyma.12G236500 overexpression transgenic positive seedlings, which can be used for phenotype analysis.
[0097] Five, salt tolerance phenotype analysis of soybean plants
[0098] (1) When the plants grow for about 3 weeks (the length of the hairy roots is 5-10 cm), the main roots are cut off, and the complex plants are transplanted into 1 / 2 Hoagland nutrient solution for recovery culture;
[0099] (2) The initial root length is measured after 2 days, then the transgenic hairy roots are placed in 120 mM NaCl solution, the growth state is observed after 6 days, and the relative root elongation rate is counted to analyze the salt tolerance phenotype. Three biological repeats are performed. Figure 3 It can be known that, under the condition of water treatment, the average relative root elongation rate of the hairy roots transformed with the empty vector (EV) is 73.33%, the average relative root elongation rate of the soybean hairy roots overexpressing Glyma.12G236500 is 73.62%, and there is no significant difference between the two, and there is no obvious difference in the growth state of the two chimeric plants; under the condition of 120 mM NaCl treatment, the average relative root elongation rate of the hairy roots transformed with the empty vector (EV) is about 1.59%, the average relative root elongation rate of the soybean hairy roots overexpressing Glyma.12G236500 is about 4.06%, and there is a significant difference between the two Figure 3 ), at the same time, the survival rate is counted, the survival rate of the soybean transformed with the empty vector (EV) is 57%, the survival rate of the soybean overexpressing Glyma.12G236500 is 70%, and there is a significant difference between the two Figure 4 ), and the soybean chimeric plants overexpressing Glyma.12G236500 are greener and grow better Figure 5 ), which shows that overexpression of Glyma.12G236500 can significantly improve the salt tolerance of soybean.
[0100] In summary, the soybean miRNA-target gene function module gma-miR482e / Glyma.12G236500 described in the application can regulate the salt tolerance of soybean, has important theoretical value and practical significance for cultivating high-quality and high-yield soybean germplasm, and has a broad application prospect in plant molecular breeding.
[0101] The above only describes the preferred embodiments of the application and is not used to limit the application. For those skilled in the art, the application can have various modifications and changes. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the application shall be included in the protection scope of the application.
Claims
1. A soybean synergistic functional module, characterized in that, The module consists of soybean microRNA gma-miR482e and its target gene Glyma.12G236500; gma-miR482 negatively regulates the activity of the target gene Glyma.12G236500, the nucleotide sequence of gma-miR482 is shown in SEQ ID NO.1, and the nucleotide sequence of the target gene Glyma.12G236500 is shown in SEQ ID NO.
2.
2. The application of the synergistic functional module of claim 1 in regulating the salt tolerance of leguminous plants, characterized in that, Salt-tolerant legumes can be obtained by overexpressing the Glyma.12G236500 gene or by inhibiting the expression or activity of gma-miR482e.
3. The application according to claim 2, characterized in that, The legume in question is soybean.
4. The application according to claim 3, characterized in that, Improved salt tolerance was manifested in the following ways: under salt stress, compared with the empty vector control, soybean chimeric plants overexpressing Glyma.12G236500 had higher leaf chlorophyll content, significantly increased relative root elongation, and better growth status.
5. A method for improving the salt tolerance of plants, characterized in that, The method is selected from any of the following: (1) By overexpressing the Glyma.12G236500 gene in the target plant, plants with higher salt tolerance than the target plant were obtained; (2) By inhibiting the expression or activity of gma-miR482e in the target plant, its negative regulation on the Glyma.12G236500 gene is relieved, and plants with higher salt tolerance than the target plant are obtained. The nucleotide sequence of gma-miR482e is shown in SEQ ID NO.1, and the nucleotide sequence of Glyma.12G236500 is shown in SEQ ID NO.
2.
6. The method according to claim 5, characterized in that, The target plant is soybean.
7. The method according to claim 5, characterized in that, The overexpression of the Glyma.12G236500 gene in method (1) is achieved by introducing the Glyma.12G236500 gene into the target plant.
8. The method according to claim 7, characterized in that, The introduction method employs Agrobacterium-mediated genetic transformation, including genetic transformation of soybean hairy roots or genetic transformation of the whole soybean plant.
9. The method according to claim 5, characterized in that, The methods for inhibiting gma-miR482e expression or activity in method (2) are selected from: introducing gma-miR482e antagonists, constructing silencing vectors, CRISPR / Cas9-mediated gene editing or methylation modification.