Medicago sativa MsKTI3 gene and application of medicago sativa MsKTI3 gene in improvement of plant salt tolerance

Abstract

The invention discloses a medicago sativa MsKTI3 gene and application of the medicago sativa MsKTI3 gene in improving the salt tolerance of plants, a candidate gene MsKTI3 related to the salt tolerance of medicago sativa is screened out through transcriptome sequencing analysis, the MsKTI3 is cloned, the medicago sativa and arabidopsis thaliana are over-expressed, and the functions of the gene are identified. According to the invention, a foundation is laid for deeply researching functions and a salt-tolerant regulation mechanism of MsKTI3, and a candidate gene resource is also provided for improving salt-tolerant molecules of alfalfa.

Description

Technical Field

[0001] The invention relates to the technical field of plant genetic engineering, and in particular to an alfalfa MsKTI3 gene and an application thereof in improving plant salt tolerance. Background Art

[0002] Alfalfa (Medicago sativa) is a high-quality perennial legume forage grass. It is widely used as a feed additive due to its high yield, rich nutritional value and good palatability. This forage grass not only promotes livestock growth and improves the quality of livestock products, but also improves the production and reproduction performance of livestock. Therefore, it occupies an important position in animal husbandry production. However, the global arable land area is constantly decreasing, and the problem of soil salinization is becoming increasingly serious, resulting in a decrease in the yield per unit area of ​​alfalfa. The accumulation of salt in the soil deteriorates the growth environment of plants. Excessive salt can cause osmotic stress, hinder the water absorption of plant roots, and cause physiological drought. At the same time, salt stress can also cause plant nutritional imbalance, inhibit photosynthesis and respiration, thereby affecting crop yields and destroying the stability of the ecosystem. In addition, salt stress can interfere with the normal metabolism of plants and produce a large amount of reactive oxygen species, such as superoxide anions and hydrogen peroxide, in plant cells. If these reactive oxygen species are not removed in time, they will cause oxidative damage to biological macromolecules such as proteins, nucleic acids and lipids in plant cells, further destroy the structure and function of cells, and bring serious survival challenges to plants. At present, my country mainly relies on imports for forage, and the import volume of alfalfa in my country has increased year by year, which has greatly restricted the development of my country's alfalfa industry. Therefore, it is of great significance to explore the genetic resources related to salt tolerance of alfalfa and explore the functions of alfalfa salt tolerance genes in the process of salt tolerance stress response for the genetic improvement of alfalfa.

[0003] Protease inhibitors are an important class of bioactive substances that regulate protein metabolism and participate in a variety of physiological and pathological processes by inhibiting protease activity. Protease inhibitors in nature are mainly divided into four categories: cysteine ​​protease inhibitors, serine protease inhibitors, metalloproteinase inhibitors and aspartic acid protease inhibitors. Serine protease inhibitors can be divided into several families according to different conserved domains, including soybean trypsin inhibitor Bowman-Birk family, potato protease inhibitor family I and II, RT I / MT II family, pumpkin inhibitor family, soybean trypsin inhibitor family (Kunitz type) and Serpin family. Among them, the Bowman-Birk family and potato protease inhibitor family I / II have been studied more, while the research on Kunitz trypsin inhibitor (KTI) in abiotic stress is relatively small. Kunitz trypsin inhibitor is a globular protein mainly found in plant storage organs, which can inhibit trypsin, chymotrypsin and serine (pro) enzymes similar to elastase. These inhibitors react rapidly with serine proteases to form stable complexes. Kunitz trypsin inhibitor has a molecular weight of about 20KDa and contains about 180 amino acid residues. It has a highly conserved three-dimensional structure, including a distinct hydrophobic core, three pairs of highly conserved disulfide bridges, a three-stranded β-fold, an N-terminal 310 helix and a C-terminal α-helix. The high conservation of this structure enables Kunitz trypsin inhibitor to specifically bind to and inhibit the activity of a variety of serine proteases. Kunitz trypsin inhibitor was first discovered and proposed by Kunitz in bovine pancreas in 1947, and later extracted from soybeans. So far, no research on the function of MsKTI3 in alfalfa has been reported, and its mechanism of regulating alfalfa tolerance is still unclear. Summary of the invention

[0004] In view of the above-mentioned deficiencies in the prior art, the object of the present invention is to provide an alfalfa MsKTI3 gene and its application in improving plant salt tolerance.

[0005] To achieve the above object, the present invention provides the following technical solutions: In a first aspect, the present invention provides an alfalfa MsKTI3 gene, the nucleotide sequence of the MsKTI3 gene is shown as SEQ ID NO.1.

[0006] Furthermore, the amino acid sequence encoded by the MsKTI3 gene is shown in SEQ ID NO.2.

[0007] In a second aspect, the present invention provides a recombinant vector containing the alfalfa MsKTI3 gene.

[0008] Furthermore, the recombinant vector is a vector for overexpressing the alfalfa MsKTI3 gene.

[0009] In a third aspect, the present invention provides a recombinant strain or a recombinant cell containing the alfalfa MsKTI3 gene.

[0010] In a fourth aspect, the present invention provides the use of the alfalfa MsKTI3 gene in improving the salt tolerance of plants.

[0011] Furthermore, the above application is the use of overexpressing alfalfa MsKTI3 gene in enhancing plant salt tolerance.

[0012] Furthermore, the plant is alfalfa or Arabidopsis thaliana.

[0013] Based on the above technical solution, the embodiments of the present invention can at least produce the following technical effects: The invention provides an alfalfa MsKTI3 gene and an application thereof in improving plant salt tolerance. The candidate gene MsKTI3 for alfalfa salt stress response is screened out through transcriptome sequencing analysis, MsKTI3 is cloned, alfalfa and Arabidopsis are overexpressed and the function of the gene is identified, which lays a foundation for in-depth research on the function and salt tolerance regulation mechanism of MsKTI3 and provides candidate gene resources for molecular improvement of alfalfa salt tolerance. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the structures shown in these drawings without paying creative work.

[0015] Figure 1 This is the electrophoresis of the PCR amplification product of the coding region sequence of MsKTI3 gene; Figure 2 It is the positive identification of the Arabidopsis strains overexpressing the MsKTI3 gene; A: PCR detection electrophoresis analysis of the Arabidopsis strains overexpressing the MsKTI3 gene, +: positive control, -: negative control, =: negative plants, 1-12: positive strains; B: Fluorescence quantitative PCR analysis of wild-type Arabidopsis and strains overexpressing the MsKTI3 gene; Figure 3Figure 1 shows the germination and growth of wild-type (WT) Arabidopsis and Arabidopsis overexpressing MsKTI3 on ​​medium with different concentrations of sodium chloride. AD: Comparison of germination and growth of wild-type (WT) Arabidopsis and Arabidopsis overexpressing MsKTI3 gene, the NaCl concentration was set to 0 mmol•L -1 (A), 100mmol•L -1 (B), 150mmol•L -1 (C) and 200mmol•L -1 (D); EH: Wild-type (WT) Arabidopsis and Arabidopsis overexpressing MsKTI3 gene at 0mmol•L -1 (E), 100mmol•L -1 (F), 150mmol•L -1 (G) and 200mmol•L -1 (H) Germination rate at different time points on sodium chloride medium; WT: wild-type Arabidopsis, At-OE2, At-OE3, At-OE9: Arabidopsis lines overexpressing the MsKTI3 gene; Figure 4 is the phenotype comparison between wild type and transgenic Arabidopsis thaliana overexpressing MsKTI3 after salt stress; AB: phenotype before NaCl stress; CD: 0mmol•L -1 (C), 250mmol•L -1 (D) Phenotypes of wild-type and overexpressing Arabidopsis plants after 17 days of NaCl stress treatment; Figure 5 The rosette leaf number and chlorophyll content analysis of wild-type and MsKTI3-overexpressing transgenic Arabidopsis lines after 17 days of salt stress treatment; A: rosette leaf number analysis chart, B: chlorophyll content analysis chart; Figure 6 This is the detection of the degree of oxidative damage of Arabidopsis leaves under salt stress; A: DAB staining; B: NBT staining, the darker the color, the more severe the degree of oxidative damage; Figure 7 Analysis of the salt tolerance phenotype of plants overexpressing the MsKTI3 gene in alfalfa roots; A: Phenotype of plants growing normally under hydroponic conditions; B: Phenotype of plants growing under 200mmol•L hydroponic conditions -1 Phenotypes after 4 days of NaCl stress, C: CAT activity detection of plants; WT: wild-type alfalfa plants, K: alfalfa plants overexpressing the MsKTI3 gene in roots. DETAILED DESCRIPTION

[0016] The technical solutions in the embodiments of the present invention will be described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. In addition, the technical solutions between the various embodiments can be combined with each other, but they must be based on the ability of ordinary technicians in this field to implement them. When the combination of technical solutions is contradictory or cannot be implemented, it should be considered that such a combination of technical solutions does not exist and is not within the scope of protection required by the present invention.

[0017] 1. Experimental Materials The alfalfa Zhongmu 4 (M. sativa cv. Zhongmu-4) and Arabidopsis thaliana (Col-0, WT) used in this experiment were provided by the forage breeding and cultivation team of the Beijing Institute of Animal Husbandry and Veterinary Medicine, Chinese Academy of Agricultural Sciences. The experimental materials were cultivated in two ways: greenhouse soil culture and artificial climate chamber. The cultivation conditions of the greenhouse soil culture method were: 22 / 20℃ (day / night), 50% humidity, 14h light / 10h dark; the cultivation conditions of the artificial climate chamber were 24 / 22℃ (day / night), 60% humidity, 16h light / 8h dark.

[0018] 2. Experimental Methods 1. Cloning and bioinformatics analysis of MsKTI3 gene from alfalfa According to the previous transcriptome data of alfalfa salt tolerance in the laboratory, the MsKTI3 gene that responds to salt stress was screened, and the full-length amplification primers of the MsKTI3 gene were designed using DNAMAN software, as shown in Table 1. According to the operating instructions of the RNA extraction kit, the RNA of the alfalfa root was extracted and reverse transcribed into cDNA. The reverse transcription system was 5×All-in-one qRT SuperMix 4 µL, Enzyme Mix 1uL, template RNA Total RNA: 1µg, RNase-Free ddH2O was added to make up to 20µL, shaken and mixed, and then centrifuged instantly to collect to the bottom of the tube. After PCR incubation at 50℃ for 15 min, heating at 85℃ for 5s, the cDNA template was synthesized and stored in a -80℃ refrigerator for later use. The CDS sequence of MsKTI3 gene was amplified using cDNA as template. The reaction system was 25μL: 2×Phanta MaX Master Mix 12.5µL, MsKTI3-F 1µL, MsKTI3-R 1µL, cDNA 2µL, RNase-FreeddH2O 8.5µL; reaction program: 98℃ 30s, 98℃ 10s, 52℃ 5s, 72℃ 4s (28 cycles), 72℃ 5min. After the length of the amplified product and the gene were correctly aligned, the gel with the target fragment was cut out using a UV gel cutter and placed in a 1.5mL centrifuge tube for gel recovery. The gel recovery product was cloned into the vector pEASY®-T5 and sequenced after the bacterial liquid PCR test was positive.

[0019] Table 1 List of primers for this experiment 2. Construction of MsKTI3 gene overexpression vector and acquisition of transgenic Arabidopsis Using restriction enzyme QuickCut TM SmaI was used to cut the pSmRY-YFP vector, and the cloning primer Y-MsKTI3-F / R containing the homology arm of the pSmRY-YFP vector was designed. The root cDNA of alfalfa No. 4 was used as a template, and the full-length coding region sequence of the MsKTI3 gene was amplified with a high-fidelity enzyme. The MsKTI3 gene fragment and the pSmRY-YFP vector that had been cut with enzymes were connected with T4 DNA ligase and transformed into DH5α Escherichia coli competent cells. The positive single colony was selected and activated after shaking. The plasmid was extracted after sequencing and alignment. The recombinant expression vector was then transformed into GV3101 Agrobacterium competent cells, and the wild-type Arabidopsis was transformed by the floral dip method. The cells were cultured in 4 mg•L -1 Homozygous plants of the T3 generation were selected on the 1 / 2MS solid medium containing glufosinate ammonium. Transgenic Arabidopsis thaliana was identified using PCR, qRT-PCR and other techniques.

[0020] 3. Analysis of salt tolerance in Arabidopsis thaliana overexpressing MsKTI3 Wild-type Arabidopsis thaliana (Col-0) and harvested T3 transgenic seeds were sterilized with 75% alcohol for 5 min, then rinsed with sterile water 4-5 times in a clean bench, vernalized at 4°C for 2 days, and then moved to a light incubator for culture and subjected to different treatments. -1 、100mmol•L -1 、150mmol•L -1 、200mmol•L -1 The seeds were cultured on 1 / 2MS solid medium containing NaCl, and the seed germination rate was counted after 10 days.

[0021] To further determine the effect of MsKTI3 gene on salt tolerance of Arabidopsis, Arabidopsis cultured in the light incubator for 14 days was transplanted to a nutrient medium with a volume ratio of 1:1 between nutrient soil and vermiculite for 17 days, and then continued with subsequent treatments. -1 NaCl solution was used for stress. Specifically, 30 ml was evenly poured into each pot with a 5 mL syringe, once every 3 days, and the phenotype was observed and physiological indicators were tested after 17 days of treatment. The determination method is as follows: relative conductivity was measured, and chlorophyll content was determined by 95% anhydrous ethanol extraction. NBT and DAB staining were performed on Arabidopsis leaves after 17 days of stress. The staining method is as follows: take leaves in the same position, wash them with distilled water, soak the samples in DAB and NBT staining solution, stain them for several hours under light-proof and room temperature conditions, then soak the samples in anhydrous ethanol, boil them in a water bath for 30 minutes, shake them intermittently during the period, and finally move the samples to filter paper soaked with 60% glycerol for photography and observation.

[0022] 4. Analysis of root growth and salt tolerance of MsKTI3 overexpressing alfalfa The amplified MsKTI3 gene fragment was connected to the linear vector pSmRY-YFP for transformation. After double identification by bacterial liquid PCR and company sequencing, the positive single clone plasmid was transformed into Ar.1193 competent cells. After sequencing analysis, the clones with correct sequences were selected and re-inoculated on TY solid medium (tryptone 5g•L -1 , yeast extract 3g•L -1 , Calcium chloride 0.6g•L -1 , agar powder 6g•L -1) streak culture. Seed germination was performed before streaking. Zhongmu No. 4 seeds were placed in a 50 mL centrifuge tube and disinfected with 75% alcohol for 8 minutes. They were washed with ddH2O 5 times and finally transferred to a 10% sodium hypochlorite solution for 15 minutes. After washing with ddH2O 5-6 times, they were placed in a 9 mm glass dish containing filter paper. The filter paper should be completely soaked and a small amount of water should be left in the dish. After that, they were wrapped in tin foil and placed in a 4°C refrigerator for 2 days. Then they were transferred to a 25°C light incubator (16h light / 8h dark) for 2 days. At this time, the seedlings grew two cotyledons and the hypocotyl was about 1-1.5 cm. The root tips of the alfalfa seedlings were cut off by 3-5 mm. After the incision was gently dipped in the bacteria on the TY plate, they were transferred to the modified Fahraes medium for co-culture. The infected seedlings were then cultured vertically in Fahraeus medium at 22°C for 7 days with 16 h light / 8 h dark. After one week, the roots growing near the radicle were completely cut off, leaving the swollen part at the bottom of the radicle. The plants with secondary root cuttings were then transferred to the selection medium (mFahraeus + 2 mg•L -1 Phosphinothricin (PPT) was used for vertical culture at 22°C for 10 days with 16 h light / 8 h dark. After 10 days, the agar on the roots of the seedlings was carefully and thoroughly washed off under running water. After 12 days of culture with 1 / 2 Hoagland nutrient solution, the positive hairy root lines were verified by PCR amplification. -1 After 4 days of NaCl stress, the phenotypes were observed and the CAT activities were measured.

[0023] 3. Results and Analysis 1. Cloning of MsKTI3 gene The MsKTI3 gene was cloned from the cDNA of alfalfa root by PCR amplification using MsKTI3-F and MsKTI3-R as primers. Figure 1 The nucleotide sequence was compared by Blast on NCBI, and the similarity with KTI3 of Medicago truncatula was 97%. The nucleotide sequence of MsKTI3 gene is shown in SEQ ID NO.1, and the amino acid sequence encoded by it is shown in SEQ ID NO.2.

[0024] SEQ ID NO.1: ATGCTAGCATTTCTTCTTCTCTTTGCATTGTCTTCGCAACCACTACTTGGATCAACTGAAGCTTCACCCGATCAAGTCATTGACACATCGGGCAAGAAGCTTCGAGCTGATACCAATTACTACATCATTCCGGCTAAGCCATTTACAATATGTGGCTTTGTTAGTTGTTTCAATAGTGGAGGCATTGCTCTTGAAACCGTCGGAGAATCTTGCCCTCTTGATGTAGTAGTTGTGAAACATAATCAAGGTTTACCGTTGAGGTTCACACCCGTTAACAACAAGAAAGGCGTTATTCGTGTCTCCACTGATCTCAACATAAAGTTCTCTAATGATGCTTATGATTCTAGATGTCCTAACCATTCCTTAGTGTGGAAGATTGATCCTTTTTCTAAAGAAGAAACATTTGTGACTACTAATGGTGTATTGGGAAACCCGGGTTCAAATACAATCCACAATTGGTTCCAAATTGAGAAGTATGAAGATGCTTATAAGTTGGTCTATTGTCCTAATGTGTGCCCTACTTGCAACCATGTATGCAAAGATATTGGAATTTACGTGTATAAGACTAGGGAAATGCGTTTGGCTCTCACTAATGTTCCCTTTAAAGTTAAGTTTCAGAAGGTTTGA。

[0025] SEQ ID NO.2: MLAFLLLFALSSQPLLGSTEASPDQVIDTSGKKLRADTNYYIIPAKPFTICGFVSCFNSGGIALETVGESCPLDVVVVKHNQGLPLRFTPVNNKKGVIRVSTDLNIKFSNDAYDSRCPNHSLVWKIDPFSKEETFVTTNGVLGNPGSNTIHNWFQIEKYEDAYKLVYCPNVCPTCNHVCKDIGIYVYKTREMRLALTNVPFKVKFQKV*。

[0026] 2. Identification and Expression Level Detection of MsKTI3 Transgenic Arabidopsis thaliana Positive Seedlings Agrobacterium tumefaciens (GV3101) containing the MsKTI3 gene was transformed into wild-type Arabidopsis thaliana (Col-0, WT) by inflorescence infection. -1 ) were screened to obtain T3 generation homozygous seeds, and a total of 12 transgenic lines were screened, such as Figure 2 As shown in A. Three transgenic lines (At-OE2, At-OE3 and At-OE9) were randomly selected, and the expression level of MsKTI3 gene in wild-type and transgenic Arabidopsis was detected by qRT-PCR. The results showed that MsKTI3 gene was successfully expressed in transgenic plants, while its expression level was not detected in wild-type Arabidopsis. Figure 2 As shown in B.

[0027] 3. Analysis of germination rate of Arabidopsis thaliana under salt stress Wild-type Arabidopsis and overexpression lines (At-OE2, At-OE3, and At-OE9) were treated with 0 mmo•L -1 、100mmo•L -1 、150mmo•L -1 and 200mmo•L -1 The germination results showed that under 0mmo•L -1 In the control group, there was no significant difference in germination rate between wild type and overexpressed Arabidopsis. -1 Under NaCl stress, there was no significant difference in germination rate between the overexpression strain and the wild type; however, under 150mmo•L -1 Under NaCl stress, the germination rate of the overexpression strain was higher than that of the wild type. -1 Under NaCl stress, Arabidopsis thaliana began to germinate from the second day. On the sixth day, the germination rates of the overexpression strains and the wild type reached the highest, and the germination rate of the overexpression strains was higher than that of the wild type. The germination rates of the strains were ranked as At-OE3>At-OE9>At-OE2>WT. Figure 3 shown.

[0028] 4. Overexpression of MsKTI3 gene promotes salt tolerance of Arabidopsis seedlings in soil In order to further determine the salt tolerance of transgenic Arabidopsis seedlings, the overexpression plants and wild-type plants were transplanted into soil (vermiculite: nutrient soil = 1:1) and grown for 3 weeks. At this time, there was no significant difference in the size of the overexpression plants and the wild-type plants. Figure 4 As shown, there was no significant difference in leaf color and leaf size between unstressed wild-type plants and transgenic plants.

[0029] Use 250mmol•L -1After NaCl stress, the growth of wild-type and transgenic lines was significantly inhibited, as shown by short plants, wilting leaves and partial yellowing, but the inhibition of transgenic lines was less severe. Figure 5 As shown in the figure, the number of rosette leaves of overexpressed Arabidopsis thaliana treated with salt stress was significantly higher than that of wild-type plants. The relevant physiological indicators of Arabidopsis thaliana under salt stress were measured and found that at 0mmol•L -1 Under NaCl stress, there was no significant difference in chlorophyll content between the overexpression strain and the wild-type strain. -1 Under NaCl stress, the chlorophyll content of the overexpression strain was significantly higher than that of the wild type (p<0.05). The above results indicate that the leaves of the overexpression strain were less damaged and had stronger photosynthesis and osmotic regulation capabilities.

[0030] 5. Analysis of the antioxidant capacity of Arabidopsis thaliana under salt stress by overexpressing MsKTI3 In the natural environment, when plants encounter salt stress, the high concentration of salt environment will cause serious oxidative damage to the plants. -1 ) was treated with Arabidopsis thaliana for 17 days, and leaves from the same position of wild-type plants and overexpression plants were taken for NBT (blue) and DAB (brown) staining tests. The results showed that under normal culture conditions, there was no obvious difference in color between the wild-type (WT) and overexpression (OE) Arabidopsis leaves; under salt stress conditions, compared with the wild-type, the overexpression strain Arabidopsis leaves showed less blue and brown, such as Figure 6 This indicates that less peroxides accumulated in the leaves of Arabidopsis thaliana overexpressing MsKTI3 and the degree of leaf oxidative damage was relatively lower.

[0031] 6. Analysis of phenotypes and physiological indicators of alfalfa overexpressing MsKTI3 in roots under salt stress The recombinant expression vector was transferred into Ar.1193 competent cells. After the colony PCR identification was correct, a single clone was picked and streaked on TY solid medium for culture. The sterilized Zhongmu No. 4 seeds were placed on moist filter paper for germination. After 7 days, the seedlings grew two cotyledons and cut the roots in a clean bench. Then, the roots containing the wound were dipped in Agrobacterium and co-cultured on the culture medium. After 7 days of culture, the cut part began to swell and grow hairy roots. The roots grown from the swollen part were cut for the second time (root cutting at this time is to prevent the occurrence of false positives, but the swollen part at the bottom of the radicle of the plant cannot be cut off), and the seedlings were transferred to the selection medium for further culture. After 7 days, hairy roots can be observed growing from the incision of the seedlings. After continuing to culture on the selection medium for 10 days, they were transferred to 1 / 2 Hoagland hydroponic nutrient solution for culture for 10 days. Transgenic positive hairy root plants were obtained by PCR identification and 200mmol·L-1 After 4 days of NaCl stress, it was found that compared with alfalfa plants overexpressing MsKTI3 in the roots, the leaves of wild-type plants turned yellow and wilted more significantly, and their growth was more obviously inhibited. The CAT activity of wild-type plants and overexpressed plants was measured, and the results showed that there was no significant difference in CAT activity between wild-type plants and overexpressed plants without salt stress. -1 After NaCl stress, the CAT activity of the overexpression plants was significantly higher than that of the wild-type plants (p<0.05). Figure 7 The above results showed that under salt stress, compared with the wild type, overexpression of MsKTI3 in the roots enabled alfalfa to maintain a relatively stronger antioxidant capacity, which ensured the normal growth of the plant to a certain extent.

[0032] The above shows and describes the basic principles and main features of the present invention and the advantages of the present invention. It should be understood by those skilled in the art that the present invention is not limited to the above embodiments, and the above embodiments and descriptions are only for explaining the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention may have various changes and improvements, and these changes and improvements fall within the scope of the present invention to be protected. The scope of the present invention to be protected is defined by the attached claims and their equivalents.

Claims

1. A gene of MsKTI3 from alfalfa, characterized in that: The nucleotide sequence of the MsKTI3 gene is shown in SEQ ID NO.

1.

2. The alfalfa MsKTI3 gene, characterized in that: The amino acid sequence encoded by the MsKTI3 gene is shown in SEQ ID NO.

2.

3. A recombinant vector containing the alfalfa MsKTI3 gene according to claim 1.

4. The recombinant vector according to claim 3, characterized in that The recombinant vector is a vector for overexpressing the alfalfa MsKTI3 gene.

5. A recombinant strain or recombinant cell containing the alfalfa MsKTI3 gene according to claim 1.

6. Use of the alfalfa MsKTI3 gene according to claim 1 in improving plant salt tolerance.

7. The use according to claim 6, characterized in that: To overexpress the alfalfa MsKTI3 gene and apply it to enhance plant salt tolerance.

8. The use according to claim 6, characterized in that: The plant is alfalfa or Arabidopsis thaliana.

Images