CN118667833A - Salt-tolerant gene LjNHX of honeysuckle and application thereof - Google Patents

Abstract

The invention discloses a honeysuckle salt tolerance gene LjNHX and application thereof, wherein LjNHX is cloned by adopting a homologous cloning method, indexes such as germination rate, root fresh weight, relative water content and the like are analyzed through arabidopsis genetic transformation experiments and physiological and biochemical analysis of transgenic plants, the salt tolerance of a LjNHX transgenic plant line is verified, and the application of LjNHX3 reserves important gene resources for directional high-quality breeding in saline-alkali soil.

Description

A honeysuckle salt-tolerant gene LjNHX3 and its application

Technical Field

The invention relates to the field of biological genetic engineering, and in particular to a honeysuckle salt-tolerant gene LjNHX3 and an application thereof.

Background Art

my country is rich in honeysuckle germplasm resources, which are usually an important source of excellent traits such as high quality, stress resistance, disease and pest resistance, and high yield. However, honeysuckle is still relatively backward in terms of germplasm collection and preservation, identification and evaluation, and innovative utilization. Its industrial development has problems such as germplasm mixing and degradation, insufficient germplasm innovation, single breeding methods, and lagging functional gene research. With the cross-disciplinary development of molecular biology, cell biology, and biochemistry, the use of molecular breeding to selectively breed high-quality and highly resistant honeysuckle has the characteristics of short cycle and excellent effect. Therefore, it is of great significance to explore the key functional genes that regulate the synthesis of effective ingredients of honeysuckle under salt stress and analyze their mechanism of action for improving the salt tolerance of honeysuckle through molecular breeding and other technical means.

Summary of the invention

In order to solve the above technical deficiencies, the present invention provides a honeysuckle salt-tolerance gene LjNHX3 and its application.

The present invention is achieved through the following technical solutions:

The invention provides an application of a honeysuckle salt-tolerant gene LjNHX3 in improving the salt tolerance of Arabidopsis thaliana and honeysuckle. The cDNA of a leaf of biennial honeysuckle "Huajin No. 6" is used as a template to carry out full-length CDS cloning to obtain a target fragment, construct an LjNHX3 expression vector, obtain transgenic Arabidopsis thaliana by an inflorescence dip method, and analyze and identify the salt tolerance of the transgenic Arabidopsis thaliana. The nucleotide sequence of the honeysuckle salt-tolerant gene is SEQ ID NO.1, and the amino acid sequence of the honeysuckle salt-tolerant gene is SEQ ID No.2.

Preferably, the forward primer for the full-length CDS clone is TGGAGAGAACACGGGGGACTCTAGAATGGCTTTCCATTTGATCAGTTC; the reverse primer is TGAACGATCGGGGAAATTCGAGCTCTCACATCAATCCTGACCCTTCAG.

Preferably, when honeysuckle is treated with 100, 200, and 300 mM NaCl, the expression level of the gene LjNHX3 shows a decreasing trend in 3-72 h.

Preferably, the germination rates of the T3 positive strains with low, medium and high expression of the LjNHX3 gene were significantly higher than those of the Col-0 type within 12h-96h after treatment with 100mM and 150mM NaCl.

Preferably, the root length of LjNHX3 transgenic Arabidopsis thaliana and Col-0 type strains were significantly reduced under 100 mM NaCl treatment, and the reduction in root length of Col-0 type was greater.

Preferably, compared with Col-0 type, the Na ⁺ content and Na ⁺ /K ⁺ ratio of roots and shoots of the LjNHX3 transgenic Arabidopsis thaliana under salt stress are significantly reduced.

Preferably, the rosette leaf wilting due to water loss in LjNHX3 transgenic Arabidopsis thaliana occurs later than in Col-0 type Arabidopsis thaliana, and the number of withered leaves is less than that in Col-0 type Arabidopsis thaliana.

Preferably, after 14 days of salt stress, the fresh weight of LjNHX3 transgenic Arabidopsis is higher than that of Col-0 Arabidopsis.

Preferably, after 14 days of salt stress, the relative water content of LjNHX3 transgenic Arabidopsis is higher than that of Col-0 Arabidopsis.

Preferably, after 14 days of salt stress, the relative conductivity of the three transgenic lines overexpressing LjNHX3 Arabidopsis thaliana was significantly reduced compared with that of Col-0 Arabidopsis thaliana.

The beneficial effects of the present invention are as follows: the present invention adopts homologous cloning method to successfully clone LjNHX3 from salt-tolerant honeysuckle varieties, and through Arabidopsis genetic transformation experiments and various analyses, reserves important gene resources for directional high-quality breeding in saline-alkali land. The present invention is of great significance for improving the salt tolerance of honeysuckle by molecular breeding and other technical means.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 shows the expression level of Lonicera japonica LjNHX3 under salt stress, where * indicates P<0.05; ** indicates P<0.01; *** indicates P<0.001; **** indicates P<0.0001.

Figure 2 shows the germination rate of LjNHX3-transgenic Arabidopsis.

Figure 3 shows the germination (A) and cotyledon greening (B) of LjNHX3-transgenic Arabidopsis.

Figure 4 shows the root length (A) and relative change rate of root length (B) of LjNHX3-transgenic Arabidopsis, where * indicates P<0.05; ** indicates P<0.01; *** indicates P<0.001; **** indicates P<0.0001.

Figure 5 shows the root length of LjNHX3-transgenic Arabidopsis.

FIG6 is an analysis of ion content in LjNHX3-transformed Arabidopsis, where * indicates P<0.05; ** indicates P<0.01; *** indicates P<0.001; **** indicates P<0.0001.

FIG. 7 is a phenotypic analysis of LjNHX3-transgenic Arabidopsis thaliana.

FIG8 is an analysis of physiological indicators of LjNHX3-transgenic Arabidopsis.

DETAILED DESCRIPTION

Example 1 Cloning and analysis of the honeysuckle salt tolerance gene LjNHX3

1 Experimental Materials and Methods

1.1 Experimental Materials

1.1.1 Plant materials, strains and carriers

1.1.1.1 Plant materials

(1) Honeysuckle (Lonicera japonica): The test material was the biennial honeysuckle "Huajin No. 6", which was planted in the medicinal plant garden of Shandong University of Traditional Chinese Medicine and identified by Professor Li Jia of Shandong University of Traditional Chinese Medicine;

(2) Arabidopsis thaliana: The test material is wild-type Arabidopsis thaliana (Columbia-0ecotype, Col-0);

1.1.1.2 Bacterial strains

(1) Escherichia coli competent cells DH5α were purchased from Takala Company;

(2) Agrobacterium GV3101 competent cells were purchased from Shanghai Weidi Company.

1.1.1.3 Carrier

The plant overexpression vector pGFPGUSplus is preserved by the research group.

1.1.2 Reagents and instruments

1.1.2.1 Reagents

Phusion high-fidelity DNA polymerase (Thermo Scientific, F530L), FastDigest XbaI (Thermo Scientific, FD0684), FastDigest SacI (Thermo Scientific, FD1133), FastDigest NcoI (Thermo Scientific, FD0573), GUS staining solution (Solarbio, G3061); LB medium, YEB medium, MS medium (Coolber, PM1110), sucrose, agar powder, silwet l-77, TritonX100, Kanamycin, Rifampicin, Hygromycin, 50×TAE, agarose, nucleic acid dye, 6×loading buffer, DL2000, D15000

1.1.2.2 Kit

FastPure Plant Total RNA Isolation Kit (Vazyme), PrimeScript RT reagent Kit with gDNAEraser (TaKaRa), Plasmid Mini Extraction Kit (Tiangen), FastPure Gel DNA Extraction Mini Kit (Vazyme).

1.1.2.3 Instruments

PCR instrument (BIO-RAD, T100), fluorescence quantitative PCR instrument (BIO-RAD, CFX96), ultra-micro-volume UV spectrometer (Nano Drop 2000), gel imager (GE, Amersham Imager600), high-speed refrigerated centrifuge (ThermoScientific), electrophoresis instrument, UV spectrophotometer, balance, high-pressure sterilizer, constant temperature oscillator, light incubator, shaker, metal bath, water bath, portable pH meter, and clean bench.

1.1.3 Preparation of culture medium

1.1.3.1 Escherichia coli culture medium

(1) Liquid LB: LB + Kanamycin 50 mg/L

(2) Solid LB: LB + agar powder 15g/L + Kanamycin 50mg/L

1.1.3.2 Agrobacterium culture medium

(1) Liquid YEB: YEB+Kanamycin 50mg/L+Rifampicin 20mg/L

(2) Solid YEB: YEB + agar powder 15g/L + Kanamycin 50mg/L + Rifampicin 20mg/L

1.1.3.3 MS medium for seed screening

1/2MS (pH 5.8): MS 4.61743g/L + sucrose 10g/L + agar powder 7.5g/L + Hygromycin B 0.8uL/1mL

1.2 Experimental methods

1.2.1 Primer design

The primers were designed by Primer Premier 6 software and synthesized by Shanghai Boshang Biotechnology Co., Ltd.

The primer sequences are shown in Tables 1 and 2.

Table 1 Cloning primers (with adapter):

Table 2 Fluorescence quantitative PCR primers:

1.2.2 Extraction of total RNA from Lonicera japonica and reverse transcription to synthesize cDNA

In early October 2021, leaves of the biennial honeysuckle variety "Huajin No. 6" were taken, quickly sealed in liquid nitrogen, and stored at -80°C. Total RNA was extracted according to the instructions of the plant RNA extraction kit (FastPure Plant Total RNA Isolation Kit), and honeysuckle cDNA was obtained according to the instructions of the reverse transcription kit (PrimeScript RT reagent Kit with gDNAEraser) for subsequent full-length CDS cloning.

1.2.3 Cloning of full-length CDS

The full-length CDS was cloned using honeysuckle cDNA as a template. The PCR reaction system is shown in Table 3 and the PCR program is shown in Table 4. The PCR product was detected by 1% agarose gel, and the band of the correct size was excised and recovered using an agarose gel recovery kit (FastPure Gel DNA Extraction Mini Kit).

Table 3 PCR system for cloning target fragments

Components Addition amount cDNA/DNA 2uL Primer R 2.5uL Primer F 2.5uL 2.5mM dNTPs 4uL Phusion DNAase 0.5uL HF Buffer 10uL dd _H2O 28.5uL Total 50uL

Table 4 PCR reaction program for insert fragment cloning

1.2.4 Construction of expression vector

1.2.4.1 Obtaining and Enzyme Digestion Verification of pGFPGUSplus Plasmid

Take out the stored pGFPGUSplus plasmid from the -20℃ refrigerator and transform it into E. coli DH5α competent cells. The specific operation is as follows:

(1) Thaw DH5α competent cells on ice and mix gently;

(2) Add 1uL of plasmid and flick to mix (if it is a recombinant plasmid, add 10uL);

(3) Place on ice for 30 min;

(4) 42°C water bath for 90 seconds;

(5) On ice for 5 min, add 500uL;

(6) 37℃, 200rpm, 45-60min;

(7) Take 50uL of bacterial solution and spread it on LB solid medium (Kan ⁺ );

(8) Incubate the cells upside down in a 37°C incubator overnight.

In the clean bench, single clones were picked and placed in liquid LB medium (Kan ⁺ ) at 37°C, 200 rpm, and shaken overnight. The pGFPGUSplus plasmid was extracted using a plasmid extraction kit, and single and double enzyme digestion verifications were performed.

(1) Single enzyme digestion verification: The plasmid was digested with NcoI enzyme and verified by 1% agarose gel. The sizes of the single enzyme digestion bands were 12000 bp, 1095 bp, and 560 bp, respectively. The plasmid was digested with XhoI enzyme and verified by 1% agarose gel. The sizes of the single enzyme digestion bands were 12600 bp and 1100 bp, respectively.

(2) Double enzyme digestion verification: XbaI+SacI enzymes were used to perform double enzyme digestion of the plasmid, and the double enzyme digestion band sizes were 12925 bp and 776 bp, respectively, as verified by 1% agarose gel.

1.2.4.2 Linearization of pGFPGUSplus vector

(1) Plant overexpression vector: Use XbaI and SacI double-enzyme digestion vector, and the double-enzyme digestion system is shown in Table 5. After the system is configured, gently blow and mix, and the digestion condition is 37°C for 2h. The digestion product is verified by 1% agarose gel, and the large fragment is purified and recovered using a gel recovery kit, which is the linearized pGFPGUSplus overexpression vector.

Table 5 Enzyme digestion system of plant overexpression vector

1.2.4.3 Homologous recombination

The linearized vector and the inserted fragment obtained above were subjected to homologous recombination reaction respectively. The reaction system is shown in Table 6. The recombination condition is 37° C. for 30 min.

Table 6 Homologous recombination reaction system

Components Addition amount Carrier 2uL Insert 2uL 5×CEⅡBuffer 4uL ExnaseⅡ 2uL _ddH2O 10uL Total 20uL

1.2.4.4 E. coli transformation and verification

The recombinant plasmid was transformed into E. coli competent cells DH5α. Single clones were picked on the second day. The correct single clones were verified by colony PCR. The bacteria were shaken at 37°C and 200 rpm overnight and sent to the company for sequencing. The reaction system and reaction procedure of colony PCR are shown in Table 7 and Table 8:

Table 7 Colony PCR reaction system

Components Addition amount Bacterial liquid 1uL Primer R 1uL Primer F 1uL 2×Mix 10uL dd _H2O 7uL Total 20uL

Table 8 Colony PCR reaction procedure

program temperature time Initial Denaturation 94℃ 4min Denaturation 94℃ 30s Annealing 55℃ 30s Extension 72℃ 90s Final extension min 72℃ 5min Hold 4℃ ∞

1.2.4.5 Chemical transformation of Agrobacterium and PCR verification

Secondary activation of the correct bacterial solution, extraction of plasmids, chemical transformation of Agrobacterium GV3101, the transformation method is as follows:

(1) Agrobacterium GV3101 was thawed on ice;

(2) Add 5uL of the recombinant plasmid and gently stir the bottom of the tube to mix;

(3) Place on ice for 5 min, place in liquid nitrogen for 5 min, place in a 37°C water bath for 5 min, and place in an ice bath for 5 min;

(4) Add 700uL of YEB liquid medium without antibiotics;

(5) 28°C, 200 rpm, shaking for 2 h;

(6) Centrifuge at 6000 r/min for 1 min, take 50 μL of the supernatant, blow and resuspend the cells, and apply YEB solid culture medium;

(7) Culture inverted in a 28°C incubator for 2 days;

(8) Single clones were selected for colony PCR verification, and positive clones were added to YEB liquid culture medium and incubated at 28°C, 200 rpm, and then stored in a refrigerator at 4°C for subsequent Arabidopsis infection.

1.2.5 Obtaining transgenic Arabidopsis thaliana by inflorescence dip method

1.2.5.1 Bacterial enrichment

(1) Activate 50 mL of bacterial solution and shake overnight;

(2) Centrifugation at 6000 rpm for 10 min to enrich the cells;

(3) Resuspend the bacteria in 5% sucrose solution, which is the infection solution.

1.2.5.2 Flower infection

(1) Select Arabidopsis plants with good growth and cut off the siliques and open flowers of Arabidopsis;

(2) Infect the flowers with the infection solution and incubate them in a moisturizing environment for 1 day;

(3) Harvest mature seeds after about one month.

1.2.6 Resistance screening and molecular detection

1.2.6.1 Seed sterilization and culture medium resistance screening

The seeds were sterilized with 75% ethanol and evenly spread on 1/2MS solid culture medium (Hyg), vernalized at 4°C for 3 days, and then cultured in a tissue culture room at 25°C with 16h light/8h darkness.

1.2.6.2 Molecular Detection

a. RNA extraction and reverse transcription: When the positive seedlings grow to the bolting stage, cut a leaf from each plant to extract RNA and reverse transcribe;

b. qRT-PCR detection of gene expression in Arabidopsis: Col-0 type Arabidopsis was used as the control.

1.2.6.3 Obtaining homozygotes

Each positive seedling was identified as a line, and T1 generation seeds were collected after maturity; three lines with low, medium and high expression levels were selected for further screening until T3 generation homozygotes were harvested for salt tolerance analysis and promoter GUS activity analysis.

1.2.7 Analysis of germination rate and root length of transgenic Arabidopsis

a. Germination rate: T3 seeds were sterilized and sown in 1/2MS medium containing 0, 50, 100, and 150 mM NaCl. Col-0 Arabidopsis was used as a control. After vernalization at 4°C for 3 days, they were cultured in a tissue culture room. The seed germination rates were recorded and photographed at 12, 24, 36, 48, 72, 84, and 96 h, respectively.

b. Root length: T3 seeds were sterilized, sown in 1/2MS medium, vernalized at 4°C for 3 days, cultured vertically at 90° in a tissue culture room for 3 days, and transplanted to 1/2MS medium containing 0 or 100 mM NaCl. After 7 days, photos were taken to record the root length.

1.2.8 Determination of ion content in Arabidopsis thaliana overexpressing LjNHX3

After vernalization, T3 seeds were planted in seedling trays filled with substrate and cultured in a light incubator for 30 days. They were irrigated with 300 mM NaCl solution for 48 hours, and the contents of Na+ and K+ in roots and shoots were determined by atomic absorption spectrometry.

1.2.9 Analysis of phenotypic and physiological indicators of Arabidopsis thaliana overexpressing LjNHX3

a. Phenotype: After vernalization, T3 seeds were planted in seedling trays filled with substrate and cultured in a light incubator for 30 days. They were watered with 300 mM NaCl solution and the phenotypic changes of Arabidopsis seedlings were observed and photographed 14 days later.

b. Fresh weight: After 14 days of salt stress, the whole Arabidopsis plant was taken out, rinsed with ddH ₂ O for 3 times, dried, and weighed for fresh weight (Fresh Weight, FW).

c. Relative Water Content (RWC): After 14 days of salt stress, the whole Arabidopsis plant was taken out, rinsed with ddH ₂ O for 3 times, dried, weighed fresh weight Wf, placed in ddH ₂ O until constant weight Wt; and dried in an oven until constant weight Wd. Each treatment was repeated 3 times.

RWC=[(W _f -W _d )/(W _t -W _d )]×100%

d. Relative conductivity (Electrical Leakage, EL): Take Arabidopsis thaliana treated with 300 mM NaCl solution for 14 days, add ddH ₂ O and place at room temperature for 24 hours, and measure the solution conductivity EC1. Boil at 100°C for 30 minutes, cool to room temperature, and measure the solution conductivity EC2.

EL = (EC1/EC2) x 100%.

2 Experimental results:

2.1 Expression level under NaCl stress

Two-year-old honeysuckle potted plants "Huajin No. 6" were treated with 1/2 Hoagland containing 100, 200, and 300 mM NaCl. Roots were collected at 0, 3, 6, 12, 24, 48, and 72 h after treatment for salt stress-induced expression analysis, with 0 h as the control for each group.

The results showed that under the treatment of 100, 200, and 300 mM NaCl, the expression level of LjNHX3 in Honeysuckle showed an overall downward trend from 3 to 72 h (see Figure 1), and the decrease in the expression level of LjNHX3 at most time points under the treatment of 100 and 300 mM NaCl reached a significant level (P<0.05), indicating that LjNHX3 is involved in the salt stress response of Honeysuckle and is a salt stress response gene.

2.2 Arabidopsis transformation and functional evaluation of LjNHX3

(1) Germination rate

Transgenic Arabidopsis thaliana overexpressing LjNHX3 was obtained by inflorescence dip method, and the T3 positive lines Line6, Line8 and Line12 with low, medium and high expression of LjNHX3 gene were selected for seed germination experiment under salt stress, and the results are shown in Figures 2 and 3. The germination rate of transgenic Arabidopsis thaliana overexpressing LjNHX3 after 96h of growth in 1/2MS medium was 97%-100%, which was no different from Col-0 type. The germination rates of the three lines of transgenic Arabidopsis thaliana overexpressing LjNHX3 within 12h-96h after treatment with 100mM and 150mM NaCl were significantly higher than those of Col-0 type. After treatment with 100 mM NaCl for 96 h, the germination rates of Line6, Line8 and Line12 increased by 11.22%, 13.00% and 13.00%, respectively; after treatment with 150 mM NaCl for 96 h, the germination rates of Line6, Line8 and Line12 increased by 30.20%, 28.82% and 27.17%, respectively.

(2) Root length

As shown in Figures 4 and 5, the root length of transgenic Arabidopsis overexpressing LjNHX3 on 1/2MS medium was not significantly different from that of Col-0 Arabidopsis (P<0.05). Compared with the growth on 1/2MS medium, 100mM NaCl treatment significantly reduced the root length of Col-0 and transgenic Arabidopsis lines, and the reduction in Col-0 root length was greater. Under salt stress, the root length of Col-0 Arabidopsis decreased by 61.33%, while the root length of Line6, Line8 and Line12 decreased by 31.57%, 46.00% and 39.91% respectively.

2.3 Ion content

After salt stress treatment (Figure 6), the Na ⁺ content in the roots and shoots of Col-0 and LjNHX3 transgenic Arabidopsis significantly increased (P<0.05), the K ⁺ content significantly decreased (P<0.05), and the Na ⁺ /K ⁺ ratio significantly increased (P<0.05). Compared with Col-0, the Na ⁺ content and Na ⁺ /K ⁺ ratio in the roots and shoots of LjNHX3 transgenic Arabidopsis significantly decreased under salt stress (P<0.05).

2.4 Phenotypic analysis

Figure 7 shows the phenotypic changes of Arabidopsis thaliana treated with 300 mM NaCl solution for 14 days after normal culture for 30 days. Under salt stress, the rosette leaves of Col-0 Arabidopsis gradually lost water and withered, and only the central leaves remained at 14 days, showing a yellow-green color. The rosette leaves of LjNHX3 Arabidopsis thaliana lost water and withered later than those of Col-0 Arabidopsis thaliana, and the number of withered leaves was less than that of Col-0 Arabidopsis thaliana. During the stress period, LjNHX3 Arabidopsis thaliana bolted normally, but as the salt stress time prolonged, the stems gradually withered from top to bottom.

2.5. Analysis of physiological indicators

Figure 8 shows that:

(1) There was no significant difference in FW between Col-0 and LjNHX3 transgenic Arabidopsis under normal conditions. After 14 days of salt stress, the FW of transgenic Arabidopsis was higher than that of Col-0, and the difference between transgenic Line12 and Col-0 was significant (P<0.05).

(2) There was no significant difference in RWC between Col-0 and LjNHX3 transgenic Arabidopsis under normal conditions. After 14 days of salt stress, the RWC of transgenic Arabidopsis was higher than that of Col-0, and the differences between transgenic Line8 and Line12 and Col-0 were significant (P<0.05).

(3) There was no significant difference in EL between Col-0 and LjNHX3 transgenic Arabidopsis under normal conditions. After 14 days of salt stress, the EL of the three transgenic lines overexpressing LjNHX3 Arabidopsis was significantly reduced compared with that of Col-0 Arabidopsis (P<0.05).

3 Conclusion

The experiments of the present invention confirmed that the Na ⁺ content and Na ⁺ /K ⁺ ratio of the roots and shoots of transgenic LjNHX3 Arabidopsis were significantly reduced (P<0.05), and the salt tolerance was significantly improved, indicating that the Na ⁺ /H ⁺ antiporter LjNHX3 is a positive regulatory gene of Honeysuckle in response to salt stress, which reduces the Na ⁺ content in the cytoplasm under stress and reduces ion toxicity, thereby enhancing the tolerance of Honeysuckle to salt stress.

The above is only a preferred implementation of this patent. It should be pointed out that for ordinary technicians in this technical field, several improvements and substitutions can be made without departing from the technical principles of this patent. These improvements and substitutions should also be regarded as the scope of protection of this patent.

Claims (10)

1. An application of a honeysuckle salt tolerance gene LjNHX in improving salt tolerance of arabidopsis thaliana and honeysuckle is characterized in that: CDS full-length cloning is carried out by taking cDNA of a leaf of biennial honeysuckle Hua Jin as a template, a target fragment is obtained, a LjNHX expression vector is constructed, transgenic arabidopsis is obtained by an inflorescence dip-dyeing method, and the salt tolerance of the transgenic arabidopsis is analyzed and identified; the nucleotide sequence of the honeysuckle salt-tolerant gene is SEQ ID NO.1, and the amino acid sequence of the honeysuckle salt-tolerant gene is SEQ ID NO.2.

2. The use according to claim 1, characterized in that: the forward primer of the CDS full-length clone is TGGAGAGAACACGGGGGACTCTAGAATGGCTTTCCATTTGATCAGTTC; the reverse primer is TGAACGATCGGGGAAATTCGAGCTCTCACATCAATCCTGACCCTTCAG.

3. The use according to claim 1, characterized in that: the expression level of the gene LjNHX is reduced in 3-72h under the treatment of 100, 200 and 300mM NaCl.

4. The use according to claim 1, characterized in that: the germination rate of the T3 generation positive strain with the LjNHX gene expressed in low, medium and high is obviously higher than that of the Col-0 type in 12-96 hours after 100mM and 150mM NaCl treatment.

5. The use according to claim 1, characterized in that: both LjNHX transgenic Arabidopsis and Col-0 strains showed a significant reduction in root length with 100mM NaCl, with a greater reduction in Col-0 root length.

6. The use according to claim 1, characterized in that: compared with Col-0 type, the LjNHX transgenic Arabidopsis thaliana has the advantage that the Na ⁺ content and the Na ⁺/K⁺ proportion of roots and buds of the transgenic LjNHX Arabidopsis thaliana under salt stress are obviously reduced.

7. The use according to claim 1, characterized in that: ljNHX3 transgenic Arabidopsis rosette leaves wither for a period of time after dehydration and wither than Col-0 type Arabidopsis, and the number of wither leaves is less than that of Col-0 type Arabidopsis.

8. The use according to claim 1, characterized in that: after 14d of salt stress, the fresh weight of LjNHX transgenic Arabidopsis thaliana is higher than that of Col-0 type Arabidopsis thaliana.

9. The use according to claim 1, characterized in that: after 14d of salt stress, ljNHX3 transgenic Arabidopsis was higher in relative water content than type Col-0 Arabidopsis.

10. The use according to claim 1, characterized in that: after 14d of salt stress, the relative conductivity of the 3 transgenic lines overexpressing LjNHX A.thaliana was significantly reduced compared to the Col-0 type A.thaliana.