Analysis and application of psgh1 gene and its promoter in sericea potentilla

Abstract

The present application relates to a kind of PsGSH1 gene and its promoter analysis and zinc resistance application of sericeous potentilla.The nucleotide sequence length of the gene is 1509bp, sequence is SEQ.NO.1, the amino acid sequence length of the gene coding is 502aa, sequence is SEQ.NO.2, the nucleotide sequence length of the gene promoter proPsGSH1 is 1541bp, sequence is SEQ.NO.3.The plant recombinant expression vector GV1300-PsGSH1-GFP and pBI121-proPsGSH1-GUS are constructed, and are transferred into model plant arabidopsis and tobacco, and obtain overexpression strain line.By analysis and verification, it is proved that the promoter proPsGSH1 contains zinc resistance cis-acting element and has activation activity.Under zinc stress, it is verified that PsGSH1 has zinc resistance function, and the gene can effectively alleviate the damage of excessive zinc to plant, and provides gene resources for cultivating zinc-tolerant plants.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of plant genetic engineering and molecular biology, and particularly relates to analysis of PsGSH1 gene and its promoter of Potentilla sericea. Through application of the gene, it is favorable to improve zinc tolerance of plants. BACKGROUND

[0002] Potentilla sericea is an important garden ground cover plant, which is widely used in the whole country, especially in the northeast. Due to its unique rhizome structure, Potentilla sericea has the characteristics of cold tolerance, poor soil tolerance and extensive management tolerance, and is often used as a ground cover to decorate the environment, and is also a good plant material for soil fixation and slope protection. Previous studies have shown that Potentilla sericea has the function of resisting heavy metals cadmium (Cd) and lead (Pb), but there is a lack of research on its resistance to high zinc environment. In addition, unreasonable discharge of "three wastes" of zinc (Zn) mining, smelting and galvanizing processing makes Zn often over-accumulates in soil. The Zn pollution in soil is spread through plant enrichment-food chain, and then threatens human and animal health. In order to explore the enrichment mechanism of Potentilla sericea to Zn at the molecular biology level, and then cultivate new zinc-tolerant varieties for the remediation of heavy metal contaminated soil, the inventors focused on the functional genes of key enzyme pathways of Potentilla sericea to Zn resistance.

[0003] Glutathione (GSH) as an intracellular antioxidant plays a key role in protecting cells from oxidation and stress, and has become the focus of scientific researchers in recent years. In plant bodies, the GSH thiol group forms glutathione chelate with heavy metal ions, and relevant studies have shown that the chelate can also be transported to vacuoles or plastids, cell membranes and other cells for detoxification. Glutathione synthetase 1 (GSH1), also known as glutamate-cysteine ligase (GCL), is a biosynthesis rate-limiting enzyme located at the front end of the GSH synthesis and metabolism pathway. Overexpression of GSH1 gene may promote the generation of GSH and increase the synthesis of plant chelate peptide (PC). Therefore, it has great practical significance for the remediation of soil heavy metal zinc pollution to use molecular biology and genetic engineering technology to improve the expression amount of GSH1 gene in plant bodies and obtain overexpression transgenic plants, thereby promoting the synthesis of a large amount of GSH. SUMMARY

[0004] Therefore, one object of the present application is to provide a Potentilla sericea glutathione synthetase gene PsGSH1 and its encoded protein. The gene contains a 1509 bp coding sequence, encodes 502 amino acids, and the nucleotide sequence of the gene is shown as SEQ. No. 1, and the amino acid sequence is shown as SEQ. No. 2.

[0005] The application also provides a PsGSH1 gene promoter (proPsGSH1) containing a 1541bp coding sequence, wherein the nucleotide sequence encoded by the promoter is shown as SEQ. No. 3.

[0006] The application also provides a biological material containing the PsGSH1 and the proPsGSH1, wherein the biological material is an expression vector, an Escherichia coli, an Agrobacterium or a transgenic cell line.

[0007] Further, the vector is GV1300-PsGSH1-GFP and pBI121-proPsGSH1-GUS.

[0008] The application also provides a zinc resistance analysis of the cis-acting element of the proPsGSH1 and a verification result of the activation activity.

[0009] The application also provides an application of the PsGSH1 gene or the biological material containing the gene in plant zinc resistance.

[0010] The plant in the application includes but is not limited to Arabidopsis thaliana and Potentilla fulgens.

[0011] The application also provides an application of the PsGSH1 gene or the biological material containing the gene in plant new variety breeding.

[0012] The purpose of the new variety breeding is to improve the zinc resistance of the plant. The PsGSH1 gene is involved in the zinc stress response of Potentilla fulgens, and the expression of the gene is induced by zinc stress.

[0013] Preferably, the PsGSH1 gene is transformed into an Arabidopsis thaliana plant to make the PsGSH1 gene overexpressed. More preferably, the PsGSH1 gene is transformed into the Arabidopsis thaliana plant by using the Agrobacterium-mediated method and the inflorescence infection method to obtain a PsGSH1 gene overexpression transgenic plant. More preferably, the PsGSH1 gene is constructed on a plant expression vector GV1300, the Agrobacterium is transformed, then the Arabidopsis thaliana inflorescence is immersed, and the overexpression transgenic plant is screened.

[0014] Compared with the prior art, the zinc resistance gene PsGSH1 of Potentilla fulgens provided by the application is a key glutathione synthetase gene, has a brand-new gene functionality, and through constructing a Potentilla fulgens plant overexpression vector, combining the Agrobacterium-mediated method to perform heterologous transformation on a model plant Arabidopsis thaliana, the influence of the zinc resistance function of the PsGSH1 gene on the zinc stress response of the plant is verified, which provides a powerful tool for the research on the key enzyme gene of the glutathione synthesis pathway of Potentilla fulgens and provides a valuable basis for the ecological restoration application of Potentilla fulgens. BRIEF DESCRIPTION OF DRAWINGS

[0015] Figure 1Electrophoresis map of PCR product for cloning 400bp sequence of 5' end of PsGSH1 gene in Example 1; wherein, lanes 1, 2, 3, 4, 5, 6 are PCR products, and M is DL2000 Marker.

[0016] Figure 2 Schematic diagram of alignment of 400bp sequence of 5' end of PsGSH1 gene in Example 1 with transcriptome sequencing result; wherein, the red box selected region is the intron region of 5' end of PsGSH1 gene.

[0017] Figure 3 Electrophoresis map of two FPNI-PCR products of proPsGSH1 in Example 1; wherein, lanes 1-2~9-3 are the second and third round PCR products of the first FPNI-PCR, lanes 1-5~9-6 are the second and third round PCR products of the second FPNI-PCR, M is DL2000 Marker, and the red box selected region is the gel recovery and sequencing fragment.

[0018] Figure 4 Full-length cloning of proPsGSH1 in Example 1; wherein, lanes 1, 2, 3, 4 are PCR products, and M is DL2000 Marker.

[0019] Figure 5 Predicted cis-acting elements of proPsGSH1 in Example 2.

[0020] Figure 6 Visualization of cis-acting elements of proPsGSH1 in Example 2.

[0021] Figure 7 Schematic diagram of construction of pBI121-proPsGSH1-GUS recombinant expression vector in Example 2.

[0022] Figure 8 GUS staining result after Agrobacterium-mediated transient transformation of pBI121-proPsGSH1-GUS in Nicotiana benthamiana in Example 2; wherein, CK - buffer is the negative control group, CK + empty Agrobacterium is the positive control group, and proPsGSH1:GUS promoter bacterial solution is the experimental group.

[0023] Figure 9 Electrophoresis map of PCR product for cloning PsGSH1 gene in Example 3; wherein, lanes 1, 2, 3 are PCR products, and M is DL2000 Marker.

[0024] Figure 10 Schematic diagram of construction of GV1300-PsGSH1-GFP recombinant expression vector in Example 3.

[0025] Figure 11 The results of PCR verification using PsGSH1-F / R for the leaf DNA extracted from the positive transgenic Arabidopsis in Example 4; wherein OE-1-9 are 9 transgenic lines, and M is DL2000 Marker.

[0026] Figure 12 The seed germination rates of each line of Arabidopsis under zinc stress in Example 5; wherein the four figures from left to right are the seed germination of each line of Arabidopsis under 0, 1, 2, 3 mM ZnSO4 stress, WT is wild type, and OE-3 / 5 / 6 are overexpression PsGSH1 lines.

[0027] Figure 13 The seed germination rates of each line of Arabidopsis under zinc stress in Example 5; wherein the four figures from left to right are the seed germination of each line of Arabidopsis under 0, 1, 2, 3 mM ZnSO4 stress, WT is wild type, and OE-3 / 5 / 6 are overexpression PsGSH1 lines.

[0028] Figure 14 The physiological indexes of each line of Arabidopsis under zinc stress in Example 5; wherein A is SOD activity, B is CAT activity, C is MDA content, and D is GSH content. DETAILED DESCRIPTION

[0029] The following examples are used to illustrate the present application, but are not used to limit the scope of the present application. If not specifically indicated, the examples are all according to the conventional experimental conditions, such as Zhichunning et al. (Plant Molecular Biology Research (Progress in Molecular Biology Research of Plant Tolerance to Heavy Metals), 2016), or the conditions suggested by the reagent kit manufacturing company.

[0030] Example 1 Intron search of 5' end of PsGSH1 gene of P. tenuifolia and cloning of proPsGSH1

[0031] The P. tenuifolia with good and uniform growth was used as the material, and the Plant DNA Maxi Kit reagent kit (D3488-01) of OMEGA company was used to extract genomic DNA. The nucleotide sequence of PsGSH1 was obtained from the data returned by Beijing Baimaik Biological Technology Company after sequencing and analyzing the transcriptome, and the first 400 bp sequence was selected for intron search of the 5' end of PsGSH1 gene. The primer PsGSH1-400bp-F / PsGSH1-400bp-R was designed by using SnapGene 4.2.4 to amplify the 5' end sequence of PsGSH1 gene. 20 μl of PCR product and 5 μl of 6×Loading Buffer were fully mixed and subjected to agarose gel electrophoresis (200 V, 400 mA, 12 min), and the correct gel strip was cut and recovered. Figure 1), product recovery was performed using OMEGA's Gel Extraction Kit (D2500-01) and sequencing (Jilin Kumuyang Biological Technology Co., Ltd.). The sequencing results were compared with the nucleotide sequence of PsGSH1 in the transcriptome, and it was found that there was a partial intron region at position 266bp Figure 2 ).

[0032] PsGSH1-400bp-F: 5'-CCAGTATGGTACTCCTTTCCCAGATGAGTTC-3'

[0033] PsGSH1-400bp-R: 5'-TGTTATCACCTTCCATGACTTTCTCCCACTC-3'

[0034] 20μl system: 2×Rapid Taq Master Mix 10μl, 10μM upstream and downstream primers 1μl each, DNA template 2μl, ddH2O 6μl.

[0035] PCR reaction program: 95℃ pre-denaturation 2min; 94℃ denaturation 10s; 58℃ annealing 30s; 72℃ extension 2min, 35 cycles; 72℃ 5min; 4℃ storage.

[0036] According to the principle of primer design of fusion primer nested PCR (FPNI-PCR), 6 specific primers with the same direction were designed using SnapGene 4.2.4 software to avoid the intron region. FPNI-PCR was divided into 3 rounds of PCR reaction, the first round was hot asymmetric PCR reaction with the DNA of Psammochloa latiflora as template, using the degenerate primer FP1-9 and the specific primer proPsGSH1-SP1; the second round was ordinary PCR reaction with the first round PCR product as template, using the nested specific primer FSP1 and the specific primer proPsGSH1-SP2; the third round was ordinary PCR reaction with the second round PCR product diluted 100 times as template, using the nested specific primer FSP1 and the specific primer proPsGSH1-SP3. Through the above 3 rounds of PCR reaction, the promoter fragment was gradually separated. The correct conditions were recovered by agarose gel electrophoresis Figure 3), and the single clone was sequenced. In the second FPNI-PCR, three specific primers were designed according to the sequence obtained in the first FPNI-PCR, and three rounds of PCR were performed. When the long enough promoter sequence was obtained, the full-length promoter cloning primers proPsGSH1-F / proPsGSH1-R were designed according to the spliced sequence, and the primers were connected to pMD-18T vector (TaKaRa, according to the instruction), and the complete promoter sequence was obtained by sequencing. Figure 4 ) The proPsGSH1 nucleotide sequence is shown in SEQ. No. 3.

[0037] The primers used are as follows:

[0038] proPsGSH1-SP1: 5'-TTACATCCAGAGGCAAGATAGC-3'

[0039] proPsGSH1-SP2: 5'-TTGTTTCTGCCCTGCAACATTG-3'

[0040] proPsGSH1-SP3: 5'-GGAGTACCATACTGGTTGTATACT-3'

[0041] proPsGSH1-SP4: 5'-GTGTTTACCACTGACAGCAAAACACG-3'

[0042] proPsGSH1-SP5: 5'-GCGGATTCAGGATTTTATAATGGGGTGT-3'

[0043] proPsGSH1-SP6: 5'-GCTGTAGCCCAACGTAATCTATGTGT-3'

[0044] proPsGSH1-F: 5'-CTGCATCCCACACAATGTTTATGCTCC-3'

[0045] proPsGSH1-R: 5'-CTGGGAAAGGAGTACCATACTGGTTGT-3'

[0046] FPNI-PCR first round 20 μl system: 2 x Rapid Taq Master Mix 10 μl, 10 μM SP1 / 4, FP1-9 primers each 1 μl, DNA template 2 μl, ddH2O 6 μl. Reaction procedure: 95℃ 90 s; 94℃ 10 s, 62℃ 30 s, 72℃ 2 min; 94℃ 10 s, 25℃ 2 min, 0.2℃ / s, 72℃ 2 min; 94℃ 10 s, 62℃ 30 s, 72℃ 2 min, 18 cycles; 72℃ 5 min.

[0047] FPNI-PCR second round 20 μl system: 2 x Rapid Taq Master Mix 10 μl, 10 μM SP2 / 5, FSP1 primers each 1 μl, first round PCR product 1 μl, ddH2O 7 μl. Reaction procedure: 95℃ 90 s; 94℃ 10 s, 62℃ 30 s, 72℃ 2 min, 30 cycles; 72℃ 5 min.

[0048] FPNI-PCR third round 20 μl system: 2 x Rapid Taq Master Mix 10 μl, 10 μM SP3 / 6, FSP2 primers each 1 μl, second round PCR product 1 μl, ddH2O 7 μl. Reaction procedure: 95℃ 90 s; 94℃ 10 s, 62℃ 30 s, 72℃ 2 min, 12 cycles; 72℃ 5 min.

[0049] Example 2 Analysis of proPsGSH1 cis-acting element of Potentilla fulgens and verification of activation activity

[0050] The amplified proPsGSH1 sequence was analyzed using the PlantCARE online website. Figure 5 Cis-element visualization was performed using Tbtools-II software. Figure 6 It was found that proPsGSH1 not only contains TATA-box and CAAT-box core elements, but also contains stress response elements G-box, MYB, ARE, CTCGA-motif, TGACG-motif, WUN-motif, among which G-box and MYB elements have been confirmed in related literature to have the function of regulating zinc stress after binding with transcription factors.

[0051] The vector map information of pBI121-GUS empty plasmid and the nucleotide sequence information of proPsGSH1 preserved by the laboratory of Northeast Forestry University College of Landscape Architecture were determined to select Hind III (AAGCTT) and BamH I (GGATCC) as the enzyme cutting sites. proPsGSH1-18T was used as a template to design homologous arm specific primers proPsGSH1-HindIII-F / proPsGSH1-BamHI-R. pBI121-GUS vector was double digested with Hind III and BamH I, and connected according to the instructions of the Novizan C112 homologous recombination kit. The recombinant product was transformed into E. coli competent DH5α (Weidi Biology, according to the instructions), and the plasmid was extracted after sequence determination to obtain the pBI121-proPsGSH1-GUS recombinant expression vector ( Figure 7 ). After the plasmid of the correct sequencing bacteria was extracted, it was transformed into Agrobacterium GV3101 (pSoup-p19) (Weidi Biology, according to the instructions). According to the method of Liang Tongyao for transiently transforming Nicotiana benthamiana (Establishment of Agrobacterium-mediated transient expression system in tobacco, Salvia miltiorrhiza and Prunella vulgaris, 2015), the pBI121-proPsGSH1-GUS Agrobacterium was injected and cultured for 36 h before staining. GUSstain Kit kit from Coolaber was used for staining. The staining results showed that proPsGSH1 had activation activity in tobacco ( Figure 8 ), providing a basis for further study of PsGSH1 gene.

[0052] proPsGSH1-HindIII-F: 5'-GACCATGATTACGCCAAGCTTTTATGCTCCAGTTGATTCTTCGAG-3'

[0053] proPsGSH1-BamHI-R: 5'-AAGGGACTGACCACCCGGGGATCCACTGGTTGTATACTACAACTTCTTTTCTTG-3'

[0054] Example 3 Cloning of PsGSH1 gene from Potentilla griffithii and construction of GV1300-PsGSH1-GFP vector

[0055] Potentilla griffithii with good growth condition was used as the material. RNA Plant Kit kit from Kangwei Company was used to extract total RNA. Takara PrimeScript TMRT reagent Kit to obtain cDNA. The nucleotide sequence of PsGSH1 was obtained from the data returned by sequencing and analyzing the transcriptome by Beijing Baimaik Biological Technology Co., Ltd., the open reading frame (ORF) sequence position was determined using NCBI (https: / / www.ncbi.nlm.nih.gov / orffinder / ), the gene accession number in the GenBank database was PQ810936, and the predicted Query coverage with Arabidopsis thaliana glutathione synthetase (glutamate-cysteine ligase) gene (NP_001190808.1) was 95%. The primers PsGSH1-F / PsGSH1-R for amplifying the ORF of PsGSH1 gene were designed using SnapGene 4.2.4. After agarose gel electrophoresis of the PCR product, the correct gel strip was cut off ( Figure 9

[0056] PsGSH1-F: 5'-CCAGTATGGTACTCCTTTCCCAGATGAGTTC-3'

[0057] PsGSH1-R: 5'-CTGAAGTTTCCCACCTCAGTATAGTAGCTCTT-3'

[0058] 50 μl PCR reaction system: 10 × Taq Buffer 5 μl, 2 mM dNTPs 5 μl, 25 mM MgSO4 3 μl, 10 μM upstream and downstream primers 1.5 μl each, cDNA template 2 μl, KOD-Plus-Neo 2 μl, ddH2O 30 μl.

[0059] PCR reaction program: 94°C pre-denaturation for 2 min; 94°C denaturation for 15 s; 62°C annealing for 30 s; 68°C extension for 2 min, 40 cycles; 72°C for 1 min; 4°C storage.

[0060] Take 4 μl of DNA purification product and mix with 1 μl of Beijing Quanshijin Biological Company -Blunt Zero Cloning Vector plant vector, 25°C water bath reaction for 15 min, to obtain PsGSH1-Blunt recombinant plasmid, transform E. coli DH5α, and sequence the single clone liquid.

[0061] ​​The vector map information of GV1300-GPF empty plasmid and the nucleotide sequence information of PsGSH1 gene preserved in the laboratory of College of Landscape Architecture of Northeast Forestry University were determined, and the Sal I (GTCGAC) and BamH I (GGATCC) enzyme cutting sites were selected. The PsGSH1-Blunt was used as a template, and the homologous arm specific primers PsGSH1-Sal I-F / PsGSH1-BamH I-R were designed. The GV1300-GPF vector was double digested with Sal I and BamH I, and the recombinant product was transformed into E. coli competent DH5α (Weidi Biology, according to the instruction) after ligation according to the instruction of Novizan C117 homologous recombination kit, and the plasmid was extracted after sequence determination, and the GV1300-PsGSH1-GFP recombinant expression vector was obtained. Figure 10

[0062] PsGSH1-Sal I-F: 5'-TTGATACATATGCCCGTCGACATGGTACTCCTTTCCCAGATGAGT-3'

[0063] PsGSH1-BamH I-R: 5'-GCCCTTGCTCACCATGGATCCGTATAGTAGCTCTTCAAAAACAGGGTCT-3'

[0064] Example 4: Transformation of Arabidopsis thaliana with plant expression vector GV1300-PsGSH1-GFP

[0065] (1) Infection of Arabidopsis thaliana

[0066] Firstly, the recombinant vector GV1300-PsGSH1-GFP plasmid obtained in Example 3 was transformed into Agrobacterium competent cells GV3101 (Weidi Biology, according to the instruction). Secondly, the Agrobacterium liquid with correct sequence was shaken to reach OD600 of 0.8-1.0 μg·mL -1 , and the bacterial body was collected by centrifugation at 4500 rpm for 10 min. The resuspension liquid was prepared: 10 g of sucrose, 0.1 g of MES, 4 μl of 0.5 mM 6-BA, 80 μl of Silwet-77, and 20 μl of 40 mM AS were added to 200 ml. The resuspended bacterial body was resuspended with the resuspension liquid, and the resuspended infection liquid reached OD600 of 0.8 μg·mL -1 , and the infection was completed. The above part of Arabidopsis thaliana in bloom was soaked in the infection liquid for 7 min, and the plant was covered with plastic wrap to keep the humidity, and was cultured in the dark for 48 h. After one week, the same method was repeated to infect once to ensure the success rate of infection.

[0067] (2) Selection of homozygous Arabidopsis thaliana

[0068] ​The T0 generation seeds of Arabidopsis were sterilized in a sterile table and evenly sowed on a screening medium (containing 50 mg·L - 1 Kana) and placed in an artificial culture room until the 4th complete, healthy, green cotyledon was grown, and then the cotyledon was transplanted into soil for further cultivation. The seeds were collected after maturation by separating the strains and bagging. The T2 and T3 generation seeds of Arabidopsis were screened by the same method as described above according to the strains.

[0069] (3) PCR identification of positive transgenic plants

[0070] The leaf RNA and DNA of the positive transgenic Arabidopsis were extracted, the RNA was reversely transcribed into cDNA, and PCR verification was performed using PsGSH1-F and PsGSH1-R. The results showed that the positive transgenic plants all contained the PsGSH1 gene Figure 11 , indicating that the PsGSH1 was successfully introduced into Arabidopsis.

[0071] Example 5: Analysis of the zinc resistance of overexpression PsGSH1 Arabidopsis

[0072] (1) Analysis of the zinc resistance of overexpression PsGSH1 Arabidopsis seeds

[0073] The T3 generation seeds of wild type and transgenic Arabidopsis strains (OE-3, OE-5, OE-6) were sowed in 1 / 2M medium containing 0, 1, 2, 3 mM ZnSO4. The results showed that the germination rates of each strain were relatively small under non-stress, all above 90%. With the gradual increase of the concentration of Zn treatment, the germination rate of the WT strain decreased to below 5%; in contrast, the lowest of the overexpression strain decreased to 13% Figure 12 .

[0074] (2) Analysis of the zinc resistance of overexpression PsGSH1 Arabidopsis seedlings

[0075] The wild type and transgenic Arabidopsis strain tissue culture seedlings were transplanted into soil, irrigated with 50 ml of 800 mg·L -1 ZnSO4 solution every 3 days, and three biological replicates were set for each strain. The plant growth state was recorded by taking pictures at 0, 7, 14, and 21 days of treatment. The leaves were sampled for three biological replicates for determination of zinc resistance physiological indexes at 14 days of treatment. The contents of GSH and malondialdehyde (MDA) were determined using the kit of Suzhou Gexi Si Biological Technology Co., Ltd. (GSH-G0206F, MDA-G0109F); the superoxide dismutase (SOD) activity was determined by the method of nitro blue tetrazolium (NBT); and the catalase (CAT) activity was determined by the ultraviolet spectrophotometer absorption method. The results showed that, after Zn stress treatment, compared with the wild type, the transgenic strains grew better, with greener leaves and lower mortality Figure 13), and the activities of antioxidant enzymes SOD and CAT and the content of non-enzymatic antioxidant GSH in the transgenic lines were significantly higher than those in the wild type ( Figure 14 Further analysis showed that the content of MDA, an index reflecting oxidative stress, in the transgenic lines was significantly lower than that in the wild type ( Figure 14 C). Thus, it was shown that the zinc resistance of the transgenic Arabidopsis was enhanced compared with the wild type, and the PsGSH1 gene could improve the zinc resistance of the plants through enzymatic and non-enzymatic systems.

[0076] Although the PsGSH1 gene and its application have been generally described and specific experimental operation schemes have been provided above, some modifications or improvements can be made on the basis of the present application, which is obvious to those skilled in the art. Therefore, the modifications or improvements made without deviating from the main technical route of the present application are within the scope of the present application.

Claims

1. A type of silky hairy cinquefoil ( Potentilla sericea ) PsGSH1 Genes, characterized by, The nucleotide sequence is SEQ. No. 1, and the amino acid sequence of the encoded protein is SEQ. No.

2.

2. A method based on the claim 1 PsGSH1 Gene promoter proPsGSH1 The sequence, characterized in that, The nucleotide sequence is shown as SEQ. No.

3.

3. An expression vector, characterized by, The vector comprises the P. sericea of claim 1 PsGSH1 gene.

4. An expression vector, characterized by, The vector comprises the promoter of claim 2 proPsGSH1 Sequence.

5. A cell, characterized in that, The cell is a cell containing the expression vector of claim 3 after transformation of a host cell, and the host cell is an E. coli or Agrobacterium cell.

6. A cell, comprising: The cell is a cell containing the expression vector of claim 4 after transformation of a host cell, and the host cell is an E. coli or Agrobacterium cell.

7. Use of the promoter sequence of claim 2, the expression vector of claim 4, the cell of claim 6, characterized in that, Agrobacterium-mediated method was used to introduce the proPsGSH into tobacco plants, and transgenic tobacco with transient expression proPsGSH1 of the target protein was obtained.

8. Use of the gene of claim 1, the expression vector of claim 3, and the cell of claim 5 in the preparation of anti-zinc transgenic plants and plant breeding, the plants being Arabidopsis thaliana or Potentilla sericea, and the breeding aiming to improve the anti-zinc ability of the plants.

9. Use according to claim 8, characterized in that, The gene of Potentilla fulgens was transformed into Arabidopsis thaliana by Agrobacterium-mediated method to obtain transgenic Arabidopsis over-expressing the gene. PsGSH1 PsGSH1 transgenic Arabidopsis over-expressing the gene.​

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