Application of citrus CsRPS2 gene in citrus canker

Abstract

The invention discloses application of a citrus CsRPS2 gene in citrus canker, and relates to the technical field of application of a gene in citrus canker, and the application is characterized in that the scab area of the citrus canker is reduced in a form of overexpressing the citrus CsRPS2 gene, and the attack degree of the citrus canker is reduced; the coding sequence of the citrus CsRPS2 gene is a nucleotide sequence as shown in SEQ ID No. 1. The citrus CsRPS2 gene, the overexpression vector and the agrobacterium containing the vector are matched for overexpression, the resistance of the citrus to the citrus canker can be effectively improved, that is, the overexpression vector is constructed, then the citrus is transiently converted, the canker attack degree of the obtained transgenic material can be reduced to 19.02%-48.53% of that of an existing citrus material, and the citrus canker resistance is improved. The attack degree of canker can be obviously reduced, and the area of disease spots is reduced.

Description

Technical Field

[0001] The present invention relates to the technical field of plant gene therapy, and particularly relates to the use of citrus CsRPS2 gene in citrus canker disease. Background Art

[0002] Citrus canker disease caused by Xanthomonas citrisubsp. citri (Xcc) severely restricts the development of the citrus industry. The citrus canker pathogen mainly infects organs such as the leaves, shoots, and fruits of host plants, with seedlings and young trees being the most severely affected. Infected plants show symptoms such as defoliation, withered shoots, weakened tree vigor, and fruit drop, resulting in a significant decline in fruit yield and quality. There are significant differences in the resistance of different citrus varieties to canker disease, with sweet orange (Citrus sinensis) being the most susceptible, followed by sour orange (Citrus aurantium) and pomelo (Citrus maxima). The widespread spread and severe damage of citrus canker disease have become one of the key factors restricting the development of the citrus industry.

[0003] Currently, the prevention and control of citrus canker disease mainly adopt a comprehensive prevention and control strategy with chemical control as the main method and biological control as the auxiliary method. However, chemical control methods pose a risk of environmental pollution and require a large amount of manpower and material resources; while biological control faces problems such as unstable effects and high costs. Therefore, breeding new disease-resistant varieties is considered to be the fundamental way to reduce the losses caused by canker disease. Traditional cross-breeding methods are difficult to meet the production needs due to their long cycle and low efficiency. With the rapid development of molecular biology technology, researchers have begun to conduct in-depth studies on the pathogenic mechanism of pathogens and the disease-resistant defense response of plants from the perspective of disease-resistant genetic engineering. In recent years, significant progress has been made in the research on the resistance of citrus to canker disease by gene engineering technology. For example, Chen Shanchun et al. (1996) successfully obtained Jincheng orange, Xinhui orange, and navel orange strains resistant to canker disease by transferring the antibacterial peptide D gene of Antheraea pernyi; Mendes et al. (2010) and Yang et al. (2011) found that after transferring exogenous genes NLS, Chit42, Xa21, and PthA into Bingtang orange, Ponkan, and sweet orange, the resistance to canker disease can be significantly improved; CsBZIP40 is a key transcription factor responsive to the infection of citrus canker pathogen and may regulate the resistance of citrus varieties through the salicylic acid (SA) pathway; the CsLOB1 gene, as the target protein of the canker disease effector PthA, enhances the sensitivity of citrus to canker disease; targeted knockout of the CsLOB1 promoter using the CRISPR / Cas9 technology can significantly improve the resistance of citrus to canker disease.

[0004] The products encoded by plant resistance genes (R genes) play a central role in directly or indirectly recognizing pathogen effector proteins or triggering downstream signal transduction in the plant innate immune system. The R protein superfamily is mainly classified by the presence or absence of a few structural motifs or domains, such as nucleotide binding site (NBS), leucine-rich repeat (LRR), Toll / interleukin-1 receptor (TIR), coiled-coil (CC), and transmembrane domain. The plant NBS-LRR protein family encodes nucleotide binding (NBS) and C-terminal leucine-rich repeat (LRR) domains. In Arabidopsis thaliana, multiple NBS-LRR genes have been identified as playing a role in disease resistance, including RPM1, RPS1, RPS2, RPS4, and RPS5. The tomato NBS-LRR proteins Mi-1.1 and Mi-1.2 have been shown to have dual regulatory roles in regulating host cell death; overexpression of VaRGA1 in Nicotiana benthamiana enhanced resistance to Phytophthora parasitica by activating salicylic acid (SA) signaling and the phenylpropanoid pathway; the cotton NBS-LRR-like protein GbaNA increased the resistance of plants to Verticillium wilt caused by Verticillium dahliae by activating reactive oxygen species production and ethylene signal transduction; overexpression of the rice NBS-LRR disease resistance gene OsBIHD1 led to enhanced expression of ethylene synthesis genes, thus participating in ethylene-mediated immunity; the wheat NBS-LRR protein TaRCR1 plays an important role in the defense response of plants to the necrotrophic fungal pathogen Rhizoctonia cerealis by regulating the scavenging and production of reactive oxygen species. Although the regulation of NBS-LRR proteins has been partially applied in plant disease resistance, there is no relevant research in the field of citrus canker.

[0005] In view of this, the present application is specifically proposed. Summary of the Invention

[0006] The object of the present invention is to provide a use of the citrus CsRPS2 gene in citrus canker, which reduces the lesion area of citrus canker and alleviates the incidence of citrus canker by overexpressing the citrus CsRPS2 gene, so as to solve the problem that citrus canker is difficult to improve in the prior art.

[0007] First, an embodiment of the present invention provides a use of the citrus CsRPS2 gene in citrus canker, including reducing the lesion area of citrus canker and alleviating the incidence degree of citrus canker in the form of overexpressing the citrus CsRPS2 gene;

[0008] The coding sequence of the citrus CsRPS2 gene is the nucleotide sequence shown in SEQ ID No.1.

[0009] As an alternative embodiment, the preparation method of the citrus CsRPS2 gene includes the following steps:

[0010] S1: Extract the total RNA of citrus leaves using an RNA extraction kit;

[0011] S2: Reverse transcribe the total RNA of citrus leaves into cDNA;

[0012] S3: Amplify the citrus CsRPS2 gene from the cDNA using primers and recover the citrus CsRPS2 gene.

[0013] As an alternative embodiment, the primers in S3 include OE-CsRPS2-F and OE-CsRPS2-R;

[0014] The primers OE-CsRPS2-F and OE-CsRPS2-R are the nucleotide sequences shown in SEQ ID No.2 and SEQ ID No.3 respectively.

[0015] As an alternative embodiment, it includes introducing the citrus CsRPS2 gene into citrus plants, and includes the following steps:

[0016] S01: Construction of the overexpression vector of the citrus CsRPS2 gene;

[0017] S02: Introduce the overexpression vector into Agrobacterium;

[0018] S03: Transform the Agrobacterium carrying the overexpression vector in citrus plants

[0019] As an alternative embodiment, the construction of the overexpression vector of the citrus CsRPS2 gene includes double digestion of the CsRPS2 coding sequence DNA fragment and the overexpression vector pLGNe with the restriction enzymes KpnΙ and EcoRΙ, followed by gel recovery and overnight ligation;

[0020] The ligation product is transformed into Escherichia coli DH5α, and the plasmid of the positive clone is extracted using a plasmid extraction kit to obtain the overexpression vector pLGNe-CsRPS2 of CsRPS2.

[0021] As an alternative implementation, step S02 includes introducing the overexpression vector pLGNe-CsRPS2 into Agrobacterium tumefaciens by electrotransformation.

[0022] As an alternative implementation, step S03 includes injecting the Agrobacterium liquid carrying the overexpression vector into citrus leaves.

[0023] As an alternative implementation, step S03 includes identifying the overexpression of the citrus CsRPS2 gene in citrus leaves using qRT-PCR primers.

[0024] As an alternative implementation, the qRT-PCR primers include RT-CsRPS2-F and RT-CsRPS2-R;

[0025] The primers RT-CsRPS2-F and RT-CsRPS2-R are nucleotide sequences shown in SEQ ID No.5 and SEQ ID No.6 respectively.

[0026] As an alternative implementation, the citrus is Wan Jincheng orange, and the overexpression of the citrus CsRPS2 gene can reduce the incidence of citrus canker in Wan Jincheng orange by 19.02% - 48.53%.

[0027] Compared with the prior art, the embodiments of the present invention have the following advantages and beneficial effects:

[0028] In the embodiments of the present invention, by overexpressing the citrus CsRPS2 gene in combination with an overexpression vector and Agrobacterium containing the vector, the resistance of citrus to citrus canker can be effectively improved. That is, an overexpression vector is constructed and then transiently transformed into citrus, and the incidence of canker in the obtained transgenic materials can be reduced to 19.02% - 48.53% of that of existing citrus materials, which can significantly reduce the incidence of canker and the lesion area. This has great application value for the molecular breeding of citrus for canker resistance and lays a foundation for cultivating resistant varieties. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention, and therefore should not be regarded as limiting the scope. For those of ordinary skill in the art, other related drawings can be obtained based on these drawings without creative efforts.

[0030] Figure 1Bioinformatics characteristics of the citrus CsRPS2 gene provided by the embodiments of the present invention; wherein, A is the chromosomal location of the citrus CsRPS2 gene, and bp is the base; B is the gene structure of citrus CsRPS2, and Exon is the exon; C is the conserved domain of the citrus CsRPS2 gene, NBS is the nucleotide binding site, LRR is the leucine repeat domain, and aa is the amino acid.

[0031] Figure 2 Structural diagram of the CsRPS2 plant overexpression vector provided by the embodiments of the present invention; wherein, GUS:NPTⅡ represents the β-glucuronidase gene; P35S represents the plant constitutive promoter derived from cauliflower mosaic virus; TNOS represents the terminator of the nopaline synthase gene.

[0032] Figure 3 Analysis chart of the CsRPS2 expression level in the transgenic materials provided by the embodiments of the present invention; wherein, ** indicates a highly significant difference compared with the control (P<0.01).

[0033] Figure 4 Comparison chart of the symptoms of non-transgenic materials and transgenic materials after inoculating with Xanthomonas citri subsp. citri for 10 days.

[0034] Figure 5 Statistical chart of the lesion size of the transgenic material leaves after inoculating with Xanthomonas citri subsp. citri for 10 days.

[0035] Figure 6 Statistical chart of the disease index of the transgenic material leaves after inoculating with Xanthomonas citri subsp. citri for 10 days. Detailed implementation manners

[0036] To make the objectives, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are some, but not all, of the embodiments of the present invention. Usually, the components of the embodiments of the present invention described and shown in the drawings here can be arranged and designed in various different configurations.

[0037] Therefore, the following detailed description of the embodiments of the present invention provided in the drawings is not intended to limit the scope of the claimed invention, but merely represents the selected embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the scope of protection of the present invention.

[0038] It should be noted that similar reference numerals and letters indicate similar items in the following drawings. Therefore, once an item is defined in one drawing, it does not need to be further defined and explained in subsequent drawings.

[0039] In the description of the present invention, it should be noted that the terms "first", "second", "third", etc. are only used for distinguishing descriptions and cannot be understood as indicating or implying relative importance.

[0040] Embodiment

[0041] The embodiment of the present invention provides a new option for improving the resistance of citrus to citrus canker, and provides a use of a citrus CsRPS2 gene in citrus canker. This use is to integrate a citrus NBS-LRR encoding gene into citrus through an expression vector, effectively improving the resistance of citrus to citrus canker, which has great application value for citrus breeding for resistance to citrus canker.

[0042] It should be noted that the embodiment of the present invention takes late Jincheng orange as the experimental object. In practical applications, this method can also be used to improve the resistance improvement of other citrus varieties to citrus canker, and is not limited by the embodiment of the present invention.

[0043] The embodiment of the present invention includes the following contents:

[0044] One: Bioinformatics analysis of the citrus CsRPS2 gene

[0045] The citrus CsRPS2 gene is located between 15463513 bp and 15467204 bp on chromosome 1 of citrus, contains 1 exon, encodes 929 amino acids, and contains NB-ARC, WH-DRP, LRR-8 and LRR-RPS2 functional domains ( Figure 1 ), and the citrus CsRPS2 gene is the nucleotide sequence shown in SEQ ID No.1.

[0046] Among them, the nucleotide sequence of SEQ ID No.1 (the CDS sequence of CsRPS2, from ATG to the stop codon) is shown as follows:

[0047]

[0048] II: Cloning of Citrus CsRPS2 Coding Sequence

[0049] 1. RNA Extraction and cDNA Synthesis

[0050] Total RNA from citrus (Wanjincheng) leaves was extracted using a plant total RNA extraction kit (Adlai, CAT: RN09), RNA quality was verified by agarose gel electrophoresis, and its concentration was measured by a concentration meter. cDNA was synthesized using a reverse transcription kit PrimeScript RTMaster Mix (TaKaRa, CAT: RR036A).

[0051] 2. PCR Amplification of CsRPS2 Coding Sequence

[0052] Primers OE-CsRPS2-F (SEQ ID No. 2), OE-CsRPS2-R (SEQ ID No. 3) and high-fidelity enzyme PrimeSTARMaxDNAPolymerase (TaKaRa, CAT: R045Q) were used to amplify the DNA fragment of the CsRPS2 coding sequence from citrus cDNA. The fragment length was 2790 bp. The amplified DNA fragment was sequenced to determine that it was the citrus CsRPS2 gene coding sequence (SEQ ID No. 1). Under ultraviolet light, a clean blade was used to cut the agarose gel block containing the target fragment, and the DNA fragment was recovered using a kit (BioFlux, CAT: BSC02M1).

[0053] PCR amplification program: 98°C, 5 min; 98°C, 30 s, 56°C, 30 s, 72°C, 1.5 min, 35 cycles; extension at 72°C for 10 min.

[0054] The nucleotide sequence of SEQ ID No.2 (pre-CDS cloning primer OE-CsRPS2-F, containing restriction site Kpn1) is as follows:

[0055] CGGggtaccATGGATTTTATAGGAACAATCC;

[0056] The nucleotide sequence of SEQ ID No.3 (primer OE-CsRPS2-R after CDS cloning, containing restriction site EcoR1) is as follows:

[0057] ACGCgaattcCTAGAACGCCTCAAACTTGCAG.

[0058] 3. Construction of CsRPS2 overexpression vector and transformation of Agrobacterium

[0059] 1. Construction of overexpression vector

[0060] Using the plant overexpression vector pLGNe, which contains the plant constitutive promoter P35S and the β-glucuronidase gene GUS:NPTⅡ( Figure 2 ). The DNA fragment of the CsRPS2 coding sequence and the overexpression vector pLGNe were double digested with the restriction enzymes KpnΙ and EcoRΙ (ThermoFisher), and then recovered by gel extraction and ligated overnight at 16°C. The ligation was carried out using the T4 DNA Ligase kit (Promega, CAT: M1801). The ligation product was transformed into Escherichia coli DH5α, and the plasmid of the positive clone was extracted using a plasmid extraction kit (Omega, CAT: D6942) to obtain the overexpression vector pLGNe-CsRPS2 of CsRPS2.

[0061] 2. Transformation of the overexpression vector into Agrobacterium

[0062] The constructed overexpression vector was introduced into Agrobacterium tumefaciens EHA105 by electroporation. The method is as follows: Take the frozen Agrobacterium competent cells EHA105 (50 μL) and thaw them on ice in advance; Add 2 μL of the plasmid of the overexpression vector to the competent cells, mix well by pipetting, and place on ice for 5 min; Transfer the mixture to the bottom of the pre-dried electroporation cuvette, place the electroporation cuvette in the card slot and adjust it to the correct position, adjust the electroporation device to the "Agr" gear, press the electroporation button, and check the electroporation data to ensure successful electroporation; Add 1 mL of LB liquid medium to the electroporation cuvette, mix well with a pipette, transfer it to a sterile centrifuge tube, and incubate it on a shaker at 260 r / min and 28°C for 60 min; Centrifuge the bacterial solution at 10000 r / min for 1 min, discard the supernatant, resuspend the cells with 100 μL of LB liquid medium, spread the resuspended cells after spreading, and incubate them in the dark at 28°C for 2 days; After the bacterial colonies grow, use the primers OE-CsRPS2-F (SEQ ID No. 2) and OE-CsRPS2-R (SEQ ID No. 3) to perform PCR verification on single colonies.

[0063] PCR reaction conditions: 94°C, 3 min; 94°C, 30 s, 58°C, 30 s, 72°C, 30 s, 30 cycles; 72°C, 10 min.

[0064] IV: Overexpression of CsRPS2

[0065] 1. Agrobacterium infection

[0066] Take 500 μL of the Agrobacterium bacterial solution containing the pLGNe-CsRPS2 plasmid and add it to 50 mL of liquid LB medium (containing kanamycin), and culture it at 28°C and 200 r / min until OD 600 = 0.5; Activate the bacterial solution of the canker pathogen until OD 600=0.5, mixed with Agrobacterium bacterial solution in equal proportions, centrifuged to obtain bacterial cells, and diluted with sterile water to OD 600 =0.5, inoculated the leaves of Wanjin orange by needle prick method, and cultured in an incubator at 28℃.

[0067] 2. qRT-PCR Analysis of Overexpressed Material

[0068] Total RNA (Adlai, CAT No: RN09) was extracted from transgenic materials, and cDNA was synthesized using the reverse transcription kit PrimeScriptRT Master Mix (TaKaRa, CAT No: RR036A). The expression of the target gene was detected by qRT-PCR. The detection primers were RT-CsRPS2-F (SEQ ID No. 6) and RT-CsRPS2-R (SEQ ID No. 7). -△△Ct The relative expression of CsRPS2 gene in transgenic materials was calculated by defining the water-treated sample as the reference factor, i.e., its CsRPS2 expression level was 1, and then calculating the multiple of the gene expression in the transgenic material relative to the reference factor 2 -△△Ct , which is the relative expression level. The results showed that the CsRPS2 gene was expressed at a higher level in transgenic materials than in wild-type plants (up to 4.7 times that of the control). Figure 3 .

[0069] qRT-PCR reaction conditions: 95°C, 3 min, 94°C, 10 s; 56°C, 10 s, 72°C, 10 s, 40 cycles; 72°C, 10 min.

[0070] The nucleotide sequence of SEQ ID No. 4 (RT-PCR identification primer for transgenic material RT-CsRPS2-F, designed within CDS) is as follows:

[0071] TGATTGCTCCACCTCCAAC;

[0072] The nucleotide sequence of SEQ ID No.5 (primer RT-CsRPS2-R after RT-PCR identification of transgenic material, designed within CDS) is as follows:

[0073] CCCATACCCCAAACTCCTATC.

[0074] 5: Evaluation of canker resistance of CsRPS2 overexpressing materials.

[0075] On the 10th day after inoculation with the mixture of Agrobacterium tumefaciens carrying pLGNe-CsRPS2 and Xanthomonas citri subsp. citri, the symptoms of citrus canker were observed and photographed, and the lesion size was statistically analyzed using Image J V1.47. The disease severity was divided into grades 0 - 7 according to the lesion area. Let R represent the lesion area. Grade 0 (R ≤ 0.25 mm 2 ), Grade 1 (0.25 mm 2 < R ≤ 0.5 mm 2 ), Grade 2 (0.5 mm 2 < R ≤ 0.75 mm 2 ), Grade 3 (0.75 mm 2 < R ≤ 1 mm 2 ), Grade 4 (1.0 mm 2 < R ≤ 1.25 mm 2 ), Grade 5 (1.25 mm 2 < R ≤ 1.5 mm 2 ), Grade 6 (1.5 mm 2 < R ≤ 1.75 mm 2 ), Grade 7 (R > 1.75 mm 2 ); The disease index was calculated according to the formula: DI = 100XΣ

number of lesions at each grade X corresponding grade value

[0076] The results showed that 10 days after inoculation with Xanthomonas citri subsp. citri, both the overexpressing plants and the WT plants grafted at the same time showed varying degrees of disease, and there were certain differences in the lesion size ( Figure 4 ). After statistical analysis, it was found that the lesion area of the transgenic materials was significantly smaller than that of the wild-type control, reduced to 12.08% - 57.77% of the control ( Figure 5 ). The disease index of the transgenic materials was significantly smaller than that of the wild-type control, reduced to 19.02% - 48.53% of the control ( Figure 6 ). Thus, it can be seen that overexpression of CsRPS2 can significantly reduce the lesion area of citrus bacterial canker and alleviate the disease severity of citrus canker.

[0077] Generally speaking, in the embodiments of the present invention, by cloning the coding sequence of citrus CsRPS2, constructing an overexpression vector, and then transforming citrus leaves, the disease severity of the obtained transgenic materials can be reduced to a maximum of 48.53% of the existing citrus, which can significantly alleviate the disease severity of canker and reduce the lesion area.

[0078] It should be noted that the CsRPS2 gene provided in the embodiments of the present invention can also be overexpressed through various techniques for use in anti-canker molecular breeding, or can be used together with other disease-resistant or disease-susceptible genes to synergistically conduct citrus anti-canker molecular breeding, which has great application value in citrus anti-canker breeding.

[0079] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention. It should be noted that the structures or components illustrated in the drawings are not necessarily drawn to scale, and the present invention omits the description of well-known components and processing technologies and processes to avoid unnecessarily limiting the present invention.

Claims

1. Use of a citrus CsRPS2 gene in citrus canker, characterized in that, Including reducing the lesion area of citrus canker and alleviating the incidence of citrus canker in the form of overexpressing the citrus CsRPS2 gene; The coding sequence of the citrus CsRPS2 gene is the nucleotide sequence shown in SEQ ID No.

1.

2. Use of a citrus CsRPS2 gene according to claim 1 in citrus canker, characterized in that, The preparation method of the citrus CsRPS2 gene includes the following steps: S1: Extract the total RNA of citrus leaves using an RNA extraction kit; S2: Reverse transcribe the total RNA of citrus leaves into cDNA; S3: Amplify the citrus CsRPS2 gene from the cDNA using primers and recover the citrus CsRPS2 gene.

3. Use of a citrus CsRPS2 gene according to claim 2 in citrus canker, characterized in that, The primers in S3 include OE-CsRPS2-F and OE-CsRPS2-R; The primers OE-CsRPS2-F and OE-CsRPS2-R are the nucleotide sequences shown in SEQ ID No.2 and SEQ ID No.3 respectively.

4. Use of a citrus CsRPS2 gene according to any one of claims 1-3 in citrus canker, characterized in that, Including introducing the citrus CsRPS2 gene into citrus plants, and including the following steps: S01: Construction of the overexpression vector of the citrus CsRPS2 gene; S02: Introduce the overexpression vector into Agrobacterium; S03: Transform the Agrobacterium carrying the overexpression vector in citrus plants.

5. Use of a citrus CsRPS2 gene according to claim 4 in citrus canker, characterized in that, The construction of the overexpression vector of the citrus CsRPS2 gene includes double-digesting the CsRPS2 coding sequence DNA fragment and the overexpression vector pLGNe with the restriction enzymes KpnΙ and EcoRΙ, followed by gel recovery and ligation overnight; The ligation product is transformed into Escherichia coli DH5α, and the plasmid of the positive clone is extracted using a plasmid extraction kit to obtain the overexpression vector pLGNe-CsRPS2 of CsRPS2.

6. Use of a citrus CsRPS2 gene in citrus canker according to claim 5, characterized in that, In S02, it includes introducing the overexpression vector pLGNe-CsRPS2 into Agrobacterium tumefaciens by electroporation.

7. Use of a citrus CsRPS2 gene according to claim 6 in citrus canker, characterized in that, In S03, it includes injecting the Agrobacterium liquid carrying the overexpression vector into citrus leaves.

8. Use of a citrus CsRPS2 gene according to claim 7 in citrus canker, characterized in that, In S03, it includes identifying the overexpression of the citrus CsRPS2 gene in citrus leaves using qRT-PCR primers.

9. Use of a citrus CsRPS2 gene according to claim 8 in citrus canker, characterized in that, The qRT-PCR primers include RT-CsRPS2-F and RT-CsRPS2-R; The primers RT-CsRPS2-F and RT-CsRPS2-R are the nucleotide sequences shown in SEQ ID No.4 and SEQ ID No.5 respectively.

10. Use of a citrus CsRPS2 gene according to claim 1 in citrus canker, characterized in that, The citrus is Wan Jincheng, and the overexpression of the citrus CsRPS2 gene can reduce the incidence of citrus canker in Wan Jincheng by 19.02% - 48.53%.

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