Saliphilous iris IhCHS1 gene capable of improving salt tolerance and jasmonate accumulation capacity of plants and application of saliphilous iris IhCHS1 gene

Abstract

The invention relates to a halophilous iris IhCHS1 gene for improving salt tolerance and jasmonate accumulation capability of a plant and application. The salt-loving iris IhCHS1 gene belongs to a plant chalcone synthase (CHS) family, an overexpression vector of the IhCHS1 gene is constructed and transferred into a salt-sensitive plant, and through growth phenotype, physiological analysis and metabolite analysis, it is found that under salt stress, compared with a wild type, the plant with the overexpressed IhCHS1 gene is remarkably increased in seedling root length, mature-period biomass and pedicel height, and the plant with the overexpressed IhCHS1 gene is remarkably increased in mature-period biomass and pedicel height. The content of jasmonic acid compounds is remarkably increased, so that overexpression of the IhCHS1 gene can remarkably enhance the salt tolerance of the plant and the accumulation amount of jasmonic acid substances, and the IhCHS1 gene can be used for improving the growth ability of the plant under salt stress and the synthesis ability of the jasmonic acid compounds. The halophilous iris IhCHS1 gene and the overexpression vector thereof have important application value in the field of plant salt stress resistance and plant secondary metabolism genetic engineering, and can be directly applied to creation of salt-tolerant new germplasm of iris and even other plants.

Description

Technical Field

[0001] The present invention relates to the technical field of plant genetic engineering, and in particular to a Halophilous Iris IhCHS1 gene for improving the salt tolerance and jasmonic acid accumulation ability of plants and an application thereof. Background Art

[0002] Salt-alkali land is an important land resource in my country. Its restoration, improvement and development and utilization are of great significance to ecological environment construction and sustainable agricultural development. By discovering the key genes related to halophyte tolerance to high salt stress, clarifying their mechanism of action, and transferring them into salt-sensitive plants, cultivating new salt-tolerant plant germplasm, and cultivating new salt-tolerant plant germplasm and new varieties, the ecological restoration and resource development of salt-alkali land can be effectively promoted.

[0003] Studies have shown that Iris plants have a certain resistance to salt and alkali and are good materials for ecological restoration of saline-alkali land. However, different varieties of plants can form different ecosystems. Cultivating different varieties of salt-tolerant plants can help restore and protect the biodiversity of saline-alkali land. This diversity helps to improve the stability and adaptability of the overall ecosystem. Therefore, how to cultivate different varieties of salt-tolerant plants is crucial.

[0004] Chalcone synthase (EC 2.3.1.74, CHS) is the first rate-limiting enzyme and key enzyme in the flavonoid biosynthesis pathway and belongs to the plant type III polyketide synthase family. The CHS gene was first isolated from parsley and subsequently cloned in a variety of monocots, dicots and gymnosperms. CHS is involved in the biosynthesis of secondary metabolites such as flavonoids and has multiple physiological functions in plants, such as participating in the formation of plant flower color and regulating plant pollen fertility. However, the function and mechanism of the CHS gene (IhCHS1) in regulating plant salt tolerance and jasmonic acid accumulation in the iris genus are still unclear, and there is no relevant literature report on the functional research of CHS gene regulating the synthesis of jasmonic acid substances in plants. Summary of the invention

[0005] To solve the above technical problems, the present invention obtains a IhCHS1 gene through mining and screening. Compared with other genes, overexpression of this gene can significantly improve the salt tolerance and jasmonic acid accumulation ability of plants, which is mainly reflected in the growth ability of plants under salt stress tolerance and the improvement of the synthesis ability of one or more substances among jasmonic acid, jasmonic acid derivatives and precursor 12-oxo-phytodienoic acid. It can be used in the creation process of new salt-tolerant and high jasmonic acid accumulation type germplasm of Iris and even other crops. The transformation method obtained by the present invention provides a new way to improve plant salt stress tolerance and jasmonic acid biosynthesis ability.

[0006] The first object of the present invention is to provide a method for improving plant salt tolerance and / or jasmonic acid accumulation ability, comprising the following steps:

[0007] Overexpressing a chalcone synthase encoding gene (IhCHS1) in the plant;

[0008] The amino acid sequence of the chalcone synthase is shown in SEQ ID NO.1.

[0009] Further, the plant includes Iridaceae plants and Cruciferae plants; preferably, the Iridaceae plants include Iris plants; the Cruciferae plants include Arabidopsis plants; most preferably, the plant is Arabidopsis thaliana.

[0010] Furthermore, the sequence of the chalcone synthase encoding gene is shown in SEQ ID NO.2.

[0011] The second object of the present invention is to provide a nucleic acid molecule encoding chalcone synthase for use in preparing a product that improves plant salt tolerance and / or jasmonic acid accumulation ability, wherein the amino acid sequence of the chalcone synthase is shown in SEQ ID NO.1.

[0012] Further, the improvement of plant salt tolerance and / or jasmonic acid accumulation ability includes improving the growth ability of plants under salt stress tolerance and improving the synthesis ability of one or more substances in plant jasmonic acid, jasmonic acid derivatives and precursor 12-oxy-phytodienoic acid. Salt stress is one of the most serious abiotic stresses in nature. Plants gradually form a series of adaptive mechanisms in the process of suffering from salt stress, such as regulating the absorption and transport of salt ions through hormone signal transduction pathways, maintaining the balance of salt ions in the cytoplasm, inducing antioxidant protection systems, accumulating osmotic regulating substances, reducing cell osmotic potential, etc. Jasmonic acid substances are a class of fatty acid derivatives containing the basic structure of cyclopentanone, mainly including free jasmonic acid and its derivatives and active precursor 12-oxy-phytodienoic acid and other compounds. Jasmonic acid substances are an important class of growth regulators and hormone signal molecules in plants, and play a very important role in regulating plant growth and development, promoting flower bud differentiation and flowering process, resisting abiotic stress and disease resistance. In addition, jasmonic acid substances are also widely used in fragrances and cosmetics.

[0013] Furthermore, the product contains a DNA sequence or RNA sequence (such as mRNA) encoding chalcone synthase, and the DNA sequence encoding chalcone synthase is shown in SEQ ID NO.2.

[0014] Further, the plant includes Iridaceae plants and Cruciferae plants; preferably, the Iridaceae plants include Iris plants; the Cruciferae plants include Arabidopsis plants; most preferably, the plant is Arabidopsis thaliana.

[0015] The present invention also provides a product for cultivating salt-tolerant and / or high jasmonic acid accumulation plants, wherein the product contains a nucleic acid molecule encoding a chalcone synthase, and the amino acid sequence of the chalcone synthase is shown in SEQ ID NO.1.

[0016] The third object of the present invention is to provide a method for cultivating transgenic plants, comprising the following steps: expressing a chalcone synthase encoding gene in a recipient plant to obtain the transgenic plant;

[0017] The transgenic plants include salt-tolerant and / or high jasmonic acid accumulation plants or plant models;

[0018] The amino acid sequence of the chalcone synthase is shown in SEQ ID NO.1.

[0019] Furthermore, the method specifically comprises the following steps: introducing a chalcone synthase encoding gene into a recipient plant, and cultivating until a homozygote expressing the chalcone synthase encoding gene is obtained, thereby obtaining the transgenic plant.

[0020] Further, the plant includes Iridaceae plants and Cruciferae plants; preferably, the Iridaceae plants include Iris plants; the Cruciferae plants include Arabidopsis plants; most preferably, the plant is Arabidopsis thaliana.

[0021] The fourth object of the present invention is to provide a chalcone synthase that improves the salt tolerance and / or jasmonic acid accumulation ability of plants, wherein the amino acid sequence of the chalcone synthase is shown in SEQ ID NO.1.

[0022] Furthermore, the salt tolerance and / or jasmonic acid accumulation ability includes the plant's growth ability under salt stress tolerance and the ability to synthesize one or more of jasmonic acid, jasmonic acid derivatives and the precursor substance 12-oxo-phytodienoic acid.

[0023] The fifth object of the present invention is to provide a nucleic acid molecule encoding the chalcone synthase.

[0024] Furthermore, the nucleotide sequence of the nucleic acid molecule is shown in SEQ ID NO.2.

[0025] The sixth object of the present invention is to provide a recombinant expression vector containing the nucleic acid molecule.

[0026] Furthermore, the backbone of the expression vector includes but is not limited to pBI121, and the promoter includes but is not limited to 35S.

[0027] The seventh object of the present invention is to provide a recombinant cell containing the nucleic acid molecule.

[0028] Furthermore, host cells include but are not limited to plant cells.

[0029] By means of the above scheme, the present invention has at least the following advantages:

[0030] The present invention uses the halophyte Halophilus halophilus as material, and clones the IhCHS1 gene related to the salt response of Halophilus halophilus by PCR technology. Further, the homologous recombination technology is used to construct its plant overexpression vector pBI121-IhCHS1, and the overexpression of the IhCHS1 gene in the salt-sensitive plant Arabidopsis is achieved by the method of Agrobacterium infection of inflorescence. Under salt stress, the root length, fresh weight at maturity, dry weight, flower stem height and jasmonic acid compound content of Arabidopsis seedlings overexpressing the IhCHS1 gene are significantly increased compared with wild-type Arabidopsis, indicating that the IhCHS1 gene can significantly enhance the salt tolerance and jasmonic acid substance accumulation of transgenic Arabidopsis, and can be used to improve the growth of plants under salt stress and the jasmonic acid biosynthesis ability, and has important application value in the field of plant salt resistance and plant secondary metabolism gene engineering.

[0031] The above description is only an overview of the technical solution of the present invention. In order to more clearly understand the technical means of the present invention and implement it according to the contents of the specification, the following is a description of the preferred embodiments of the present invention in conjunction with detailed drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] In order to make the contents of the present invention more clearly understood, the present invention is further described in detail below based on specific embodiments of the present invention in conjunction with the accompanying drawings.

[0033] Figure 1 is the fold change of expression of different CHS genes in Halophilous Iris after salt stress treatment;

[0034] Figure 2 This is a schematic diagram of the secondary structure of the IhCHS1 protein of the halophilous iris;

[0035] Figure 3 This is a schematic diagram of the tertiary structure of the IhCHS1 protein of the haloiris;

[0036] Figure 4 To construct the overexpression vector of IhCHS1 gene of Halophilous Iris and identify the transgenic plants by PCR.

[0037] Figure 5 Effects of salt stress on root elongation of wild-type and IhCHS1 transgenic Arabidopsis seedlings;

[0038] Figure 6 Effects of salt stress on the growth of flower stems at maturity in wild-type and IhCHS1 transgenic Arabidopsis thaliana;

[0039] Figure 7 Effects of salt stress on biomass of wild-type and transgenic IhCHS1 Arabidopsis at maturity;

[0040] Figure 8 To investigate the effect of overexpression of the IhCHS1 gene on the accumulation of jasmonic acid substances in transgenic Arabidopsis thaliana. DETAILED DESCRIPTION

[0041] The present invention is further described below in conjunction with the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it, but the embodiments are not intended to limit the present invention.

[0042] The scheme involved in the present invention is as follows:

[0043] (1) Screening of key genes: The present invention screened the gene expression profile of Iris halophila under salt stress (obtained by transcriptome sequencing) and found that compared with other genes, the expression of IhCHS1 gene was significantly upregulated under salt stress ( Figure 1 ), therefore, the IhCHS1 gene was overexpressed in subsequent transgenic experiments in order to improve the salt tolerance of plants.

[0044] (2) construct a plant overexpression vector of the IhCHS1 gene;

[0045] (3) Transforming the constructed plant overexpression vector (pBI121-IhCHS1) into plants;

[0046] (4) Molecular biological techniques were used to screen out positive transgenic plants, and physiological and metabolomics techniques were used to identify salt-tolerant plants, which significantly enhanced the plant's ability to accumulate jasmonic acid.

[0047] The sequence information involved in the embodiments of the present invention is as follows:

[0048] IhCHS1 protein sequence (SEQ ID NO.1)

[0049] MVNVAELCKLQRAEGPAAILAIGTANPPNAVSQSEYPDYYFRITNSEHKHELKAKFKRMCDNSMIKKRYMYLTEEILKENPSLCAYMAPSLDVRQDMVVVEVPKLGKEAAAKAIKEWGQPKSKITHVVFCTTSGVDMPGADYQLTRLLGLRPSVKRLMMYQQGCFAGGTVLRLAKDLAENNRGARVLVVCSEITAVTFRGPSDSHLDSLVGQALFGDGASALIVGADPIENVERPLFEIVSAAQTILPDSEGAIDGHVREAGLTFHLLKDVPGIISKNIEKSLEEAFKPLGIADWNSLFWVAHPGGPAILDQVEAKLGLKPEKLRATRHVLSEYGNMSSACVLFILDEMRKRSVEAGNGTTGEGLEWGVLFGFGPVAAA

[0050] IhCHS1 gene sequence (SEQ ID NO.2)

[0051]

[0052] Example 1 Physicochemical properties and structural analysis of Iris halophila IhCHS1

[0053] 1. Based on the amino acid sequence encoded by the IhCHS1 gene, the molecular weight (MW), theoretical isoelectric point (pI) and extinction coefficient of the IhCHS1 protein were predicted using the online programs pI / MW (http: / / web.expasy.org / compute_pi / ) and Peptide and Protein Molecular Weight Calculator (https: / / www.aatbio.com / tools / calculate-peptide-and-protein-molecular-weight-mw), and the subcellular localization of the IhCHS1 protein was predicted using the online tool Cell-PLoc 2.0 (http: / / www.csbio.sjtu.edu.cn / bioinf / Cell-PLoc-2 / ). The results showed that the IhCHS1 protein of the halophilic iris chalcone synthase contained 379 amino acids, and the predicted molecular weight (MW) of the IhCHS1 protein was 41.228 kDa, the isoelectric point (pI) was 6.28, and the extinction coefficient was 35120 M -1 cm -1 , the predicted subcellular localization is in the cytoplasm.

[0054] Further use of SignalP

[0055] (https: / / services.healthtech.dtu.dk / services / SignalP-6.0 / ) predicted the signal peptide of the amino acid sequence of IhCHS1 of Halophilous Iris. The results showed that the probability value of the signal peptide was low, indicating that the amino acid sequence of IhCHS1 of Halophilous Iris may not contain a signal peptide.

[0056] 2. The secondary structure and tertiary structure of IhCHS1 protein were predicted using the online analysis tools PSIPRED (http: / / bioinf.cs.ucl.ac.uk / psipred / ) and SWISS-MODEL (https: / / swissmodel.expasy.org / interactive), respectively.

[0057] like Figure 2As shown in Figure 3, PSIPRED prediction results show that the secondary structure of IhCHS1 protein contains 19 α helices and 8 β folds, and contains the characteristic sequence of chalcone synthase (RLMMYQQGCFAGGTVLR) and four conserved catalytic active sites (Cys 164, Phe 215, His 303 and Asn 336); in addition, there is a typical chalcone synthase receptor binding pocket in the 3D structure of IhCHS1 protein ( Figure 3 ). The above structural analysis showed that IhCHS1 protein may have the catalytic activity of chalcone synthase.

[0058] Example 2 Cloning of the IhCHS1 gene of Halophilous Iris and construction of a plant overexpression vector

[0059] 1. Immerse the seeds of Iris halophila Pall. in 5% (v / v) sodium hypochlorite for sterilization and then wash them with ddH2O 5 times. After sterilization, plant the seeds in a container filled with clean, moist sand for germination. When the seedlings reach a height of about 10 cm, they are transferred to a plastic cup containing 1 / 2 Hoagland nutrient solution for cultivation, and the light intensity of the incubator is 22,500 Lux. The seed germination and seedling growth conditions are as follows: 25 / 22°C (day / night temperature), 60-80% (relative humidity) and 14 / 10-h (day / night photoperiod). After one week of cultivation, the aboveground and root samples of the Iris halophila seedlings were collected after being treated with a nutrient solution containing 300 mM NaCl for 24 h, and then frozen in liquid nitrogen for extraction of total RNA.

[0060] 2. Use the plant total RNA extraction kit (TaKaRa, China) to extract RNA from the seedlings of Iris halophila. The operation steps were carried out according to the instructions of the kit. During the extraction process, RNase-free DNase I (TaKaRa, China) was used to remove genomic DNA. After the extracted RNA was tested and qualified, cDNA was synthesized using a reverse transcription kit (TaKaRa, China).

[0061] 3. Based on the mRNA sequence obtained by full-length transcriptome sequencing, upstream primers and downstream primers were designed for amplifying the full-length CDS of the IhCHS1 gene. The XbaⅠ sequence on the plant overexpression vector pBI121 and a 15bp-long sequence upstream of the site (5'-gagaacacgggggactctagaATGGTGAATGTTGCAGAACTATGC-3') were added to the 5' end of the upstream primer, and the SmaⅠ sequence and a 15bp-long sequence downstream of the site (5'-ataagggactgaccacccgggTCACGCAGCAGCAACCGG-3') were added to the 5' end of the downstream primer. Using the cDNA of the salt-loving iris as a template, PCR cloned the IhCHS1 gene. After the PCR product was detected by 1% agarose gel electrophoresis, it was cut and recovered. At the same time, the pBI121 plasmid was double-digested with XbaⅠ and SmaⅠ, and the digested products were recovered. Using the homologous recombinase ExnaseTMII ( The PCR product was connected with the enzyme-cut product by using ⅡOne Step Cloning Kit, Vazyme, and the connected product was transformed into Escherichia coli competent DH5a, and then sequenced. The sequencing results showed that the IhCHS1 gene fragment was successfully connected with the pBI121 restriction site, indicating that the plant overexpression vector pBI121-IhCHS1 was successfully constructed ( Figure 4 A), the target fragment sequence was determined to be SEQ ID NO.2.

[0062] Example 3 Genetic transformation of IhCHS1 of Halophilous Iris and analysis of its function in salt tolerance of Arabidopsis

[0063] 1. Inoculate Agrobacterium GV3101 containing the recombinant plasmid pBI121-IhCHS1 stored in a -80°C refrigerator into YEB solid medium (containing Kan: 50 μg / ml; Rif: 50 μg / ml), and culture in the dark at 28°C for 48 hours. Inoculate the activated Agrobacterium into 5 ml YEB liquid medium (containing Kan: 50 μg / ml; Rif: 50 μg / ml), and culture at 28°C with shaking at 250 rpm for 24 hours. Pipette 2 ml of the above bacterial solution into 200 ml YEB liquid medium (containing Kan: 50μg / ml; Rif: 50μg / ml), shake culture at 250rpm at 28℃, when its OD600 reaches 1.0-1.2, the bacterial solution is divided into 50mL centrifuge tubes, centrifuged at 4℃, 4000rpm for 10min, added 5% sucrose to resuspend, centrifuged again at 4000rpm for 10min, and the supernatant is discarded. The precipitate is the bacteria. Add 5% sucrose solution containing 0.02% surfactant (silwet L-77) to make the bacterial solution OD600 value reach about 0.8-1.0, soak the Arabidopsis inflorescence in the above Agrobacterium bacterial solution for about 1-5min, take out the Arabidopsis, use absorbent paper to absorb the bacterial solution on the stem, wrap it with plastic wrap to increase humidity, and after dark culture for 24h, remove the insurance film for routine culture, spray water 1-2 times a day to increase humidity. When the transformed Arabidopsis seed pods turned yellow, the first generation of IhCHS1 transgenic seeds were harvested, dried at 37°C, and stored in a 4°C refrigerator.

[0064] 2. Arabidopsis seeds were sown on MS solid medium containing Kan (50 μg / ml) and placed in an incubator for cultivation. The seedlings that could grow normally on the screening medium were transferred to the nutrient medium for cultivation and propagation. After four weeks, the leaves of the transgenic Arabidopsis seedlings were cut to extract genomic DNA. PCR identification was performed using Arabidopsis DNA as a template and specific primers on the vector and target gene. Plants identified as positive by PCR were used for subsequent homozygous screening and transcription level detection. By statistically analyzing the segregation ratio of kanamycin resistance traits of PCR-positive offspring plants, the T3 generation single-copy insertion homozygous IhCHS1-OE1 and IhCHS1-OE2 were finally screened for subsequent experiments.

[0065] 3. In order to determine whether the transcription of IhCHS1 gene in transgenic Arabidopsis thaliana IhCHS1-OE1 and IhCHS1-OE2 can be carried out normally, RNA of wild-type and transgenic Arabidopsis thaliana was extracted respectively, two specific primers were designed, and the transcription level was detected by fluorescence quantitative PCR.

[0066] The results showed that the expression of IhCHS1 gene was not detected in wild-type Arabidopsis, while high expression of IhCHS1 gene was detected in transgenic plants IhCHS1-OE1 and IhCHS1-OE2. The above research results indicate that the IhCHS1 gene of Halophilous Iris has been successfully overexpressed in Arabidopsis ( Figure 4 B), the next step is biological function analysis.

[0067] 4. After disinfection, the T3 homozygous and wild-type Arabidopsis seeds of the IhCHS1 gene were sown on MS medium (containing 1% sucrose and 1% agar), synchronized at 4 degrees for 3 days, and then placed in a light incubator for vertical culture. The culture conditions were: day / night temperature 22°C / 20°C, light provided by LED lights, a cycle of 16 / 8 hours (day / night), and air relative humidity of 60-80% (RH). After growing for 2 days, wild-type and IhCHS1 Arabidopsis plants with consistent growth were selected and transplanted to medium containing 100mM, 150mM and 175mM NaCl, respectively. Normal medium was used as a control, and the root length was measured after 2 weeks of growth to test the effect of overexpression of the halophilic iris chalcone synthase gene IhCHS1 on the root elongation of Arabidopsis seedlings.

[0068] like Figure 5 As shown in the figure, under normal culture and 100mM NaCl treatment conditions, there was no significant difference in the growth of transgenic IhCHS1 Arabidopsis and wild-type Arabidopsis, and there was no significant difference in the root length between the two; under high salt (150mM and 175mM NaCl) treatment conditions, compared with the control group, the growth of transgenic IhCHS1 Arabidopsis and the wild type was significantly inhibited, but the degree of inhibition was significantly different between the two. The root length of transgenic IhCHS1 Arabidopsis was significantly greater than that of the wild type. After 2 weeks of 150mM NaCl treatment, the root elongation of IhCHS1-OE1 and IhCHS1-OE2 transgenic lines was 86.66% and 114.97% higher than that of the wild type, respectively. After 2 weeks of high salt (175mM NaCl) treatment, the root elongation of IhCHS1-OE1 and IhCHS1-OE2 transgenic lines was 36.7% and 26.46% higher than that of the wild type, respectively. The above results indicate that overexpression of IhCHS1 from Halophilous Iris can significantly enhance the salt tolerance of transgenic Arabidopsis seedlings.

[0069] 5. To further understand the effect of overexpression of IhCHS1 in salt-loving iris on the salt tolerance of transgenic Arabidopsis thaliana at maturity, transgenic and wild-type seedlings grown on MS medium for 2 weeks were transferred to hydroponic conditions for cultivation. After 4 weeks, they were treated with salt (150 mM NaCl) and normal nutrient solution (1 / 4 Hogland) was used as the control. After 6 days of 150 mM NaCl treatment, the growth phenotype of the plants at maturity was photographed, and the flower stem height and biomass were measured.

[0070] The study found that under normal conditions, there was no significant difference in the growth of wild-type and transgenic IhCHS1 Arabidopsis, but after 6 days of salt treatment, the growth status of transgenic lines IhCHS1-OE1 and IhCHS1-OE2 was significantly better than that of wild-type plants ( Figure 6 A), especially the height of the flower stem of transgenic IhCHS1 Arabidopsis was significantly higher than that of wild-type plants ( Figure 6 B). In addition, under salt stress conditions, the fresh weight and dry weight of the aboveground part of transgenic IhCHS1 Arabidopsis were significantly greater than those of wild-type plants ( Figure 7 The above research results show that overexpression of IhCHS1 in Halophilous Iris can significantly enhance the salt tolerance of transgenic Arabidopsis thaliana at maturity.

[0071] Example 4 Analysis of the accumulation characteristics of jasmonic acid substances in plants transgenic for the halophilous iris IhCHS1 gene

[0072] 1. First, the T3 homozygous transgenic IhCHS1 gene and wild-type Arabidopsis seeds were sterilized and sown in MS medium (containing 1% sucrose, 1% agar), synchronized at 4 degrees for 3 days, and then placed in a light incubator for vertical culture. The culture conditions are shown in Example 3. Select 2-week-old seedlings with consistent growth and transfer them to nutrient solution (1 / 4 Hogland) for culture. After 10 days of culture, different strains were treated with salt (100mM NaCl). After 24 hours of NaCl treatment, whole plant samples of Arabidopsis of each strain were taken for qualitative and quantitative analysis of metabolites.

[0073] 2. Weigh 50 mg of Arabidopsis samples from different strains, add 1000 μL of extract containing internal standard (methanol, acetonitrile and water volume ratio = 2:2:1) for grinding and ultrasonic treatment, centrifuge at 4°C, 12000rpm for 15min, and dry the supernatant in a vacuum concentrator. The vacuum-dried extract was redissolved in acetonitrile-water solution (v / v = 1:1), and the metabolites in the samples to be tested were analyzed using an ultra-high performance liquid chromatography-tandem high-resolution mass spectrometer system. Metabolite qualitative and quantitative analysis mainly used the raw data collected by MassLynx V4.2 to perform peak extraction, peak alignment and other data processing operations through Progenesis QI software. Based on the online METLIN database and public database of Progenesis QI software, identification was performed, and theoretical fragment identification was performed at the same time, with the deviation of the parent ion mass number within 100ppm and the deviation of the fragment ion mass number within 50ppm.

[0074] 3. Further comparison of KEGG, HMDB and LIPID MAPS databases combined with metabolite quantification results was used to screen and identify jasmonic acid compounds with significant differences in accumulation in different strains.

[0075] The results showed that compared with the wild-type control, under salt stress conditions, the accumulation of most compounds in the jasmonic acid metabolic pathway in the transgenic lines overexpressing IhCHS1 showed a significant (p<0.05) increase, including jasmonic acid (Jasmonic acid and 7-epi-Jasmonic acid) and its synthetic precursor (12-OPDA) and derivatives [Methyl jasmonate and Jasmonoyl-(L)-isoleucine] ( Figure 8 ). This indicates that under salt stress, the chalcone synthase gene IhCHS1 of the halocyanine has the function of enhancing the biosynthesis of jasmonic acid substances.

[0076] Obviously, the above embodiments are merely examples for clear explanation and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived from these are still within the protection scope of the invention.

Claims

1. A method for improving plant salt tolerance and / or jasmonic acid accumulation ability, characterized in that: The following steps are involved: overexpressing a chalcone synthase encoding gene in the plant; The amino acid sequence of the chalcone synthase is shown in SEQ ID NO.

1.

2. The method according to claim 1, characterized in that Contains at least one of the following characteristics: (1) The plants include plants of the Iridaceae family; (2) The plants include plants of the family Cruciferae; (3) The plant includes an iris plant; (4) The plant includes a plant of the genus Arabidopsis; (5) The plant includes Arabidopsis thaliana.

3. Use of a nucleic acid molecule encoding a chalcone synthase in the preparation of a product that improves plant salt tolerance and / or jasmonic acid accumulation ability, characterized in that: The amino acid sequence of the chalcone synthase is shown in SEQ ID NO.

1.

4. The use according to claim 3, characterized in that: The improving of the salt tolerance and / or jasmonic acid accumulation capacity of the plant includes improving the growth capacity of the plant under salt stress and / or improving the synthesis capacity of one or more of jasmonic acid, jasmonic acid derivatives and the precursor substance 12-oxo-phytodienoic acid.

5. The use according to claim 3, characterized in that: Contains at least one of the following characteristics: (1) The product contains a DNA sequence or RNA sequence encoding chalcone synthase; (2) The plant includes a plant of the Iridaceae family and / or a plant of the Cruciferae family; preferably, the plant of the Iridaceae family includes a plant of the genus Iris; the plant of the Cruciferae family includes a plant of the genus Arabidopsis; most preferably, the plant is Arabidopsis thaliana.

6. A product for cultivating salt-tolerant and / or high jasmonic acid accumulating plants, characterized in that: The product contains a nucleic acid molecule encoding chalcone synthase, and the amino acid sequence of the chalcone synthase is shown in SEQ ID NO.

1.

7. A method for cultivating transgenic plants, characterized in that: The method comprises the following steps: expressing a chalcone synthase encoding gene in a recipient plant to obtain the transgenic plant; The transgenic plants include salt-tolerant and / or high jasmonic acid accumulation plants or plant models, The amino acid sequence of the chalcone synthase is shown in SEQ ID NO.

1.

8. The method according to claim 7, characterized in that Contains at least one of the following characteristics: (1) The method specifically comprises the following steps: introducing a chalcone synthase encoding gene into a recipient plant, cultivating until a homozygous plant expressing the chalcone synthase encoding gene is obtained, thereby obtaining the transgenic plant; (2) The plant includes a plant of the Iridaceae family and / or a plant of the Cruciferae family; preferably, the plant of the Iridaceae family includes a plant of the genus Iris; the plant of the Cruciferae family includes a plant of the genus Arabidopsis; most preferably, the plant is Arabidopsis thaliana.

9. A chalcone synthase for improving plant salt tolerance and / or jasmonic acid accumulation ability, characterized in that: The amino acid sequence of the chalcone synthase is shown in SEQ ID NO.

1.

10. A nucleic acid molecule encoding the chalcone synthase according to claim 9, a recombinant expression vector or a recombinant cell containing the chalcone synthase encoding gene.

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