Rice adiponectin receptor ospls3 protein and coding gene and application thereof

Abstract

The application discloses a rice anti-Rhizoctonia solani leptin receptor OsPLS3 protein and an encoding protein and application thereof. The rice leptin receptor OsPLS3 protein has an amino acid sequence as shown in SEQ ID NO. 3. A coding gene of the rice leptin receptor gene OsPLS3 has a genomic nucleotide sequence as shown in SEQ ID NO. 1, and an open reading frame nucleotide sequence of the rice leptin receptor gene OsPLS3 is as shown in SEQ ID NO. 2. Experiments prove that overexpression of the rice leptin receptor gene OsPLS3 in the application in rice can improve the resistance of the rice to the Rhizoctonia solani, and the rice is more susceptible to the Rhizoctonia solani after knockout, namely, the OsPLS3 positively regulates the resistance of the rice to the Rhizoctonia solani. The rice leptin receptor gene OsPLS3 has important production value for breeding of the rice resistant to the Rhizoctonia solani.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of plant genetic engineering, and particularly relates to a rice adiponectin receptor OsPLS3 protein and a coding gene and application thereof. BACKGROUND

[0002] More than 50% of the world's population takes rice as the main food crop, which is of extremely important significance for the long-term survival and development of human beings. Rice sheath blight is one of the main fungal diseases of rice, which is distributed in all major rice producing areas in the world, and seriously affects the yield and quality of rice.

[0003] Adiponectin is very important for regulating human diabetes, and adiponectin receptors can recognize adiponectin to start adiponectin-induced cell activities. In 2005, scholars searched for homologues of human adiponectin receptor AdipoR1 in Arabidopsis thaliana, and identified 5 genes, which were named as HHP (heptahelical protein) gene family due to their typical 7 transmembrane domains, including AtHHP1 (At2g24150), AtHHP2 (At4g30850), AtHHP3 (At4g37680), AtHHP4 (At4g38320) and AtHHP5 (At5g20270) (Hsieh et al., 2005, A novel gene family in Arabidopsis encoding putative heptahelical transmembrane proteins homologousto human adiponectin receptors and progestin receptors, J EXP BOT, doi: 10.1093 / jxb / eri311). Through phylogenetic evolution analysis, it was found that they all belong to the PAQRs (Progestin and adiponectin receptors) family, and the structure of AtHHPs family is similar to AdipoRs, which consists of an intracellular N-terminal domain, a 7TM domain and a shorter extracellular C-terminal fragment (Fernandes et al., 2005, Regulated expression of putative membrane progestin receptor homologues in human endometrium and gestational tissues. The Journal of endocrinology, doi: 10.1677 / joe.1.06242),

[0004] It has been reported that AtHHPs are involved in plant ABA and NaCl stress, and the germination rate of hhp1 mutant of Arabidopsis is significantly reduced compared with Col-0 after adding NaCl and ABA (Chen et al., 2009, HHP1 is involved in osmotic stress sensitivity in Arabidopsis. J EXP BOT, doi:10.1093 / jxb / erp039). At the same time, it has been found that AtHHP1 is also involved in cold stress, and the transcription factor MYB96 induces the expression of AtHHP1 / 2 / 3 / 4 after feeling cold stress, and AtHHPs activate the downstream ICE1 to improve the cold resistance of Arabidopsis (Lee et al., 2015, The MYB96-HHP module integrates cold and abscisic acid signaling to activate the CBF-COR pathway in Arabidopsis, The Plant journal: for cell and molecular biology, doi:10.1111 / tpj.12866).

[0005] The reports on HHPs gene family mainly involve abiotic stress (salt stress, cold stress and ABA signal), and whether HHPs are involved in biotic stress has never been reported, and the present application identifies 6 OsHHPs by searching the homologues of Arabidopsis AtHHPs in rice, and studies the immune function of OsHHP3 (LOC_Os03g13040) among them, and finds that the rice transgenic line overexpressing OsHHP3 can confer resistance to Rhizoctonia solani. Since HHPs belong to the PAQRs receptor family, the present application renames them as PLSs (PAQR-like sensors) according to their gene characteristics. SUMMARY

[0006] In order to solve the above technical problems, the present application provides a rice anti-Rhizoctonia solani leptin receptor OsPLS3 protein and its encoding protein and application.

[0007] The rice leptin receptor OsPLS3 protein provided by the present application has an amino acid sequence as shown in SEQ ID NO. 3.

[0008] The encoding gene of the OsPLS3 protein also belongs to the protection scope of the present application.

[0009] The cDNA sequence of the encoding gene is shown in SEQ ID NO. 2.

[0010] The expression cassette containing the coding gene, the recombinant expression vector, the transgenic recombinant bacteria or the transgenic cell line also belong to the protection scope of the present application.

[0011] The present application also provides the application of the rice adiponectin receptor OsPLS3 protein or the coding gene in regulating the disease resistance of plants.

[0012] Preferably, the plant is rice, and the disease resistance is the disease resistance to sheath blight.

[0013] Further, the present application provides the application of the rice adiponectin receptor OsPLS3 protein in the improvement of rice germplasm resources.

[0014] In the application, the method for improving the rice germplasm resources is to overexpress the coding gene of the rice adiponectin receptor OsPLS3 protein in rice, so as to enhance the resistance to rice sheath blight.

[0015] In one embodiment of the present application, the present application provides a method for cultivating a transgenic plant with improved disease resistance, which is to transfer the coding gene into a plant, and to screen a transgenic plant overexpressing the coding gene, i.e. a transgenic plant with improved disease resistance.

[0016] Preferably, the plant is rice, and the disease resistance is the disease resistance to rice sheath blight.

[0017] In another embodiment of the present application, the present application provides a method for inhibiting the disease resistance of rice, which is to reduce the activity of the rice adiponectin receptor OsPLS3 protein in rice or to completely inactivate the rice adiponectin receptor OsPLS3 protein, or to inhibit the transcription of the coding gene of the rice adiponectin receptor OsPLS3 protein or to delete the coding gene of the rice adiponectin receptor OsPLS3 protein, so as to obtain rice with reduced disease resistance; the disease resistance is the disease resistance to rice sheath blight.

[0018] Preferably, the activity of the rice adiponectin receptor OsPLS3 protein in rice is reduced or the rice adiponectin receptor OsPLS3 protein is completely inactivated by knocking out the coding gene (OsPLS3 gene) of the rice adiponectin receptor OsPLS3 protein.

[0019] In the present application, the screening and identification method of the positive strain of overexpressed coding gene (rice adiponectin receptor OsPLS3 gene) includes the following steps:

[0020] RNA of the transgenic rice is extracted, and the RNA is reversely transcribed into cDNA, and then the transcription level of the OsPLS3 gene in the wild type and the overexpressed strain is detected by real-time fluorescent quantitative PCR; if the transcription level of the OsPLS3 gene in the transgenic rice is obviously higher than that in the wild type, it is said that the transgenic strain is the positive strain of overexpressed OsPLS3, and if the transcription level of the OsPLS3 gene is not higher than that in the wild type, it is said that the transgenic strain is the negative strain of the OsPLS3 gene.

[0021] Experiments prove that the rice adiponectin receptor gene OsPLS3 in the present application can improve the disease resistance of rice to sheath blight after overexpression in rice, the length of sheath blight lesion is shortened, and transgenic plants with improved resistance to sheath blight can be obtained. Therefore, the rice adiponectin receptor in the present application has great practical application value for cultivating sheath blight-resistant rice materials.

[0022] The method for identifying the homozygous line of the knockout material of the rice OsPLS3 gene comprises the following steps:

[0023] 1) Design primers according to the nucleotide sequence of the rice adiponectin receptor gene OsPLS3 and construct an OsPLS3 gene knockout vector;

[0024] 2) Genetically transform rice with the vector to obtain a transgenic rice line

[0025] 3) The identification of the homozygous gene knockout rice line mainly comprises the following steps:

[0026] For the constructed knockout plant: design primers according to the target, extract leaf genomic DNA of the transgenic plant, amplify and purify the DNA fragment using the primers, and perform sequencing. If the sequencing result shows that a mutation occurs and is a single peak, it indicates that it is a transgenic positive homozygous plant.

[0027] The beneficial effects of the present application are:

[0028] Overexpression of the rice adiponectin receptor gene OsPLS3 in rice can confer resistance to sheath blight of rice. After inoculation with sheath blight fungus, the length of lesion of OsPLS3 transgenic plants is significantly reduced compared with wild type, and the gene can be used for disease resistance breeding. BRIEF DESCRIPTION OF DRAWINGS

[0029] Figure 1 It is the agarose gel electrophoresis result of OsPLS3 CDS amplification.

[0030] Figure 2 It is the detection of hygromycin in the genomic DNA of OsPLS3 overexpression transgenic line.

[0031] Figure 3 It is the detection of OsPLS3 gene transcription level in the leaf of OsPLS3 overexpression transgenic rice line.

[0032] Figure 4 It is the detection of hygromycin and OsPLS3 gene in the genomic DNA of ospls3 knockout line.

[0033] Figure 5 It is the sequencing result of CAS9 target site of ospls3 knockout homozygous line.

[0034] Figure 6 Results of ospls3 transgenic lines inoculated with rice sheath blight fungus in vivo, A, lesion figures of each transgenic line inoculated with rice sheath blight fungus, B, statistical results of lesion length of each line, error bars represent standard error data of the mean, using one-tailed test: ns, no significant difference; *, p < 0.005; **, p < 0.001.

[0035] Figure 7 Results of transgenic lines inoculated with rice sheath blight fungus in vitro, A, lesion figures of each transgenic line inoculated with rice sheath blight fungus, B, statistical results of lesion area of each line, error bars represent standard error data of the mean, using one-tailed test: **, p < 0.001; ***, p < 0.0001. DETAILED DESCRIPTION

[0036] Unless otherwise specifically indicated, the techniques employed are conventional techniques within the skill of the art and experimental methods were performed according to conventional methods; the materials, reagents, etc. used are commercially available.

[0037] Example 1, obtaining of rice adiponectin receptor OsPLS3 protein and its encoding gene

[0038] The cDNA sequence was obtained by reverse transcription of Zhonghua 11 RNA, and then the LOC_Os03g13040 CDS sequence was queried through the Phytozome database (https: / / phytozome-next.jgi.doe.gov / ). The genomic sequence of the gene contains 5'UTR (1-533 bp), exon 1 (533-766 bp), intron 1 (766-2542 bp), exon 2 (2542-2867 bp), intron 2 (2867-3494 bp), exon 3 (3494-4148 bp), 3'UTR (4148-4532 bp), 3 exons (SEQ ID NO. 1). The CDS sequence is 1212 DNA bases (SEQ ID NO. 2), encoding 403 amino acids (SEQ ID NO. 3).

[0039] Forward and reverse primers were designed based on the OsPLS3 CDS:

[0040] OsPLS3-F: 5'-ATGGCGGCGGCGGCGGGGG

[0041] OsPLS3-R: 3'-GCAGCCTTGCACGTCTCTCC

[0042] The PCR reaction system is as follows:

[0043] Reagents (20 μL reaction system)

[0044]

[0045] Amplified in medium flower 11 cDNA, PCR amplification results are shown in Figure 1 The amplified OsPLS3 CDS sequence of the application is proved to be completely same as the CDS sequence (SEQ ID NO. 3) in Phytozome database by company sequencing.

[0046] Example 2, rice adiponectin receptor OsPLS3 protein and its encoding gene function verification

[0047] I. Obtaining of OsPLS3 gene overexpression and knockout homozygous rice strains

[0048] 1. Construction of OsPLS3 gene overexpression vector:

[0049] The open reading frame nucleotide (SEQ ID No. 2) of rice adiponectin receptor gene OsPLS3 is selected at both ends of about 20 bp, and the upstream primer and downstream primer are designed, and the sequences are as follows:

[0050] pCAMBIA1305-OsPLS3-F:

[0051] 5'-GCAGCCCGGGGGATCCATGGCGGCGGCGGCGGGGG

[0052] pCAMBIA 1305-OsPLS3-R:

[0053] 3'-CTTTGTAGTCAAGCTTGCAGCCTTGCACGTCTCTCC

[0054] The vector used was pCAMBIA1305 (Gangling Li et al., 2022, RGN1 controls grain number and shapes panicle architecture in rice, Plant Biotechnol J, doi:10.1111 / pbi.13702), and the pCAMBIA1305 vector was digested with BamHI / HindIII. The full-length open reading frame (ORF) of the rice adiponectin receptor gene OsPLS3 (SEQ ID No. 2) was amplified using cDNA from rice Zhonghua 11 as a template. The digested vector and the gene fragment amplified by the primers were ligated using a homologous recombinase. The vector was then heat-shocked into *E. coli*, and the plasmid was extracted. Sequencing was performed using the vector primers. The recombinant expression vector containing SEQ ID No. 2, verified by sequencing, was identified as the OsPLS3 gene overexpression vector. The extracted plasmid was then heat-shocked into *Agrobacterium* EHA105. Two days later, single *Agrobacterium* clones were picked for colony PCR detection. Correct *Agrobacterium* clones were shaken, and after two days, the shaken *Agrobacterium* was transferred to preservation tubes, mixed with an equal volume of 40% glycerol, and stored at -80℃ for later use. High-success-rate PCR enzyme KOD FX (Code No. KFX-101, Toyobo (Shanghai) Biotechnology Co., Ltd.) was used during gene amplification to ensure fidelity under conditions of high GC content.

[0055] 2. Construction of gene knockout vectors

[0056] Two sgRNAs were designed using the first 20 bp of the NGG sequence in the exon of the rice adiponectin receptor gene OsPLS3 as target sites. Both sgRNA1 and sgRNA2 target the first exon.

[0057] sgRNA1: 5'-GTACGGGCTTGTGGAGTACC-3';

[0058] sgRNA2-R: 5'-AGACGTTGAGGGTCTCGTTG-3';

[0059] Then, OsPLS3-sgRNA1-F and OsPLS3-sgRNA2-R were designed as forward and reverse primers to construct the knockout vector:

[0060] OsPLS3-sgRNA1-F: 5'-TTGTGCAGATGATCCGTGGCGTACGGGCTTGTGGAGTACCGTTTTAGAGCTAGAAATA-3';

[0061] OsPLS3-sgRNA2-R: 5'-ACTTGCTATTTCTAGCTCTAAAACAGACGTTGAGGGTCTCGTTGTGAGCCTCAGCGCAG-3';

[0062] The specific construction method is as follows:

[0063] The vectors used in the process are pYLsgRNA-OsU6a, pUC19-OsU3-gRNA and pYLCRISPR / Cas9-MH (Ma et al., 2015, A Robust CRISPR / Cas9 System for Convenient, High-Efficiency Multiplex Genome Editing in Monocot and Dicot Plants, Mol Plant, doi:10.1016 / j.molp.2015.04.007, publicly available from China Agricultural University). OsPLS3-sgRNA1-F and OsPLS3-sgRNA2-R are used to amplify pYLsgRNA-OsU6a as a template:

[0064] 1) PCR reaction system

[0065] Reagents (20 μL reaction system)

[0066]

[0067] The components are added to the PCR tube, centrifuged briefly, and then placed in a PCR instrument, pre-denatured at 95°C for 3 min, denatured at 95°C for 15 s, annealed at 50-60°C for s, extended at 72°C for 1 min / Kb, and the denaturation, annealing and extension steps were performed for 35 cycles, followed by 72°C extension for 5 min, and 10°C termination of the PCR reaction program. The PCR product is electrophoresed using 1% agarose gel, recovered from the gel, and further recovered from the gel. The recovered product is ligated with the pUC19 vector using EcoRV and XbaI endonuclease to cut the pUC19-OsU3-gRNA vector:

[0068]

[0069] The gene fragment and the vector are then subjected to homologous recombination by the Novozyme kit (ClonExpress II One Step Cloning Kit, C112):

[0070]

[0071] Then the ligation product was heat-shocked into E. coli, positive clones were screened using ampicillin selection, and then positive clones were detected by colony PCR. The positive clones were shaken to extract plasmids, sequenced using vector primers, and the correct ones were obtained by LR reaction to the pYLCRISPR / Cas9-MH-GW vector (recorded in the literature "Zhao Z#, Xie X#, Liu W#, Huang J, Tan J, Yu H, Zong W, Tang J, Zhao Y, Xue Y, Chu Z, Chen L*, Liu Y-G* (2022). STIPCR: an efficient method for amplification and de novo synthesis of long DNA sequences. Molecular Plant 15(4): 620-629." which is publicly available from China Agricultural University). OsPLS3 gene knockout vector was obtained.

[0072] 3. Obtaining of transgenic lines:

[0073] The above constructed OsPLS3 gene overexpression and knockout vectors were sent to Wuhan Boyuan Biotechnology Co., Ltd. for genetic transformation, and transgenic lines were obtained about three months later.

[0074] 4. Detection of overexpression transgenic positive lines:

[0075] The obtained transgenic seedlings were taken out of the test tube and planted in nutrient soil. After two weeks of culture, leaf samples of each single plant of T0 generation were taken, frozen quickly in liquid nitrogen, and ground. Plant genomic DNA was extracted by CTAB method, and then hygromycin specific primers were used for detection. If hygromycin can be detected, it means that the vector has been transferred into the line. Figure 2 The seeds of this line were subsequently harvested, and T1 generation seeds were screened on hygromycin 1 / 2MS plates to obtain a 3:1 trait segregation. At this time, the wild type was removed, and homozygous and heterozygous transgenic plant seedlings were planted again and T2 generation seeds were harvested. Hygromycin 1 / 2MS plates were used for screening again, and the line that did not segregate was the homozygous line.

[0076] II. RNA level expression detection of OsPLS3 overexpression lines:

[0077] 1. Extraction of RNA (Trizol method):

[0078] 1) Cut 0.4g rice leaves, freeze in liquid nitrogen, and put in a grinder to break at 50Hz / min for 1min, repeat 3 times, add 1mL Trizol lysis solution, shake to form homogenate, and let stand at room temperature for 5min to fully separate the nucleic acid and protein complex. (Note: the volume of the extracted sample should not exceed one-tenth of the volume of the added Trizol lysis solution, otherwise there will be DNA contamination in the extracted sample). 2) After 5min, add 200μL of chloroform and shake thoroughly on the shaker for 20s, mix thoroughly, and let stand at room temperature for 10min after the solution becomes cloudy. 3) Centrifuge the sample at 12000rpm / min for 10min in a 4°C centrifuge, and after centrifugation, the solution will separate into three layers, carefully pipette the colorless water phase containing RNA (usually 500μL is pipetted to avoid contamination), and transfer to a new RNAse-free centrifuge tube. 4) After liquid transfer, add an equal volume of isopropanol to the centrifuge tube, shake thoroughly on the shaker for 20s, and let stand at room temperature on the bench for 10min to allow the RNA to precipitate. 5) Centrifuge the sample at 12000rpm / min for 10min in a 4°C centrifuge, at this time the white precipitate begins to appear at the bottom of the centrifuge tube, carefully pipette the upper liquid and discard. 6) Add 1mL of 75% RNAse-free alcohol, gently shake the precipitate, then centrifuge at 12000rpm / min for 10min in a 4°C centrifuge, discard the supernatant. 7) Repeat step 6). 8) Place in a sterile ultraclean bench for 30min until the ethanol is completely volatilized, then add 40μL of RNase-free water, gently blow with a pipette gun until the precipitate is completely dissolved. 9) Measure the RNA concentration with a Nanodrop and store at -80°C for future use.

[0079] 2. Reverse transcription (MF166-plus-01, Beijing Polymer Biotech Co., Ltd.):

[0080] The total reverse transcription system is 20μL, divided into two steps

[0081] 10x gDNA plus remover mix 1μL

[0082] RNA template 1μg

[0083] DEPC-H2O to 10μL

[0084] Mix and put in PCR instrument, 42°C for 2min, complete and quickly insert into ice.

[0085] Add to the PCR tube of the previous step

[0086] 5x M5 RT Super plus Mix 4μL

[0087] DEPC H2O 6 μL

[0088] After mixing gently, put into PCR instrument, 37℃ reverse 15min, 85℃ heating 5s, take out to obtain cDNA solution, use immediately or save to -20℃ refrigerator.

[0089] 3. Fluorescent quantitative PCR:

[0090] The experimental method of quantitative real time polymerase chain reaction (qRT-PCR) refers to FastSYBR Mixture (Kangwei Century Biotechnology Co., Ltd.). The primers use OsPLS3-qpcr-F: 5'-TACGGGCTTGTGGAGTACCG-3', OsPLS3-qpcr-F: 5'-TCCTATCAAGTGCGTCCAGAC-3'.

[0091] 1) PCR reaction system

[0092] Reagent (20 μL reaction system)

[0093]

[0094] 2) PCR reaction conditions

[0095] Pre-denaturation 95℃ 30s

[0096] Denaturation 95℃ 10s

[0097] Annealing / extension 60℃ 30s

[0098] The denaturation and annealing / extension steps are repeated for 40 cycles.

[0099] The results of quantification are analyzed by Bio-Rad CFX Manager, first analyze its melting curve, QC (quality control), etc. Remove substandard data, use software with Gene Study function according to (1+E) -ΔΔ Ct algorithm analyzes the expression of target genes. Figure 3 ). Figure 3 The results show that the OsPLS3 gene overexpression vector constructed by us has higher expression than the wild type after transformation.

[0100] III. Identification of OsPLS3 knockout homozygous lines

[0101] The leaves of homozygous knockout lines were quickly frozen with liquid nitrogen and ground, and the plant genomic DNA was extracted by CTAB method, and then hygromycin specific primers were used for detection. If hygromycin can be detected, it means that the knockout vector has been introduced into the strain Figure 4 ). In the figure, the negative control ZH11 did not detect hygromycin, while ospls3-1-16 could detect hygromycin, indicating that the CAS9 vector has been introduced. By designing specific primers with an amplification length of about 500 bp upstream and downstream of the knockout target, the PCR products were amplified and then electrophoresed on a 4% agarose gel, and compared with the wild type amplification products Figure 4 ), the PCR products with different band sizes from the wild type were sent to the company for sequencing. The sequencing results showed that the target position was mutated or deleted, and the sequencing results were single peak, then the strain was homozygous mutant Figure 5 , ospls3-1, ospls3-2 are two homozygous mutant lines), the strain was collected for subsequent experiments.

[0102] IV. In vivo inoculation of transgenic rice lines to verify the resistance to rice sheath blight fungus

[0103] The high temperature and high humidity climate from July to August in summer is most suitable for inoculating sheath blight fungus outdoors. Rice sheath blight fungus (Rhizoctonia solani Kühn) GD-118 (Yang et al., Cloning and functional analysis of an endo-PG-encoding gene Rrspg1 of Rhizoctonia solani, the causal agent of rice sheath blight, Canadian Journal of Plant Pathology, doi: 10.1080 / 07060661.2012.709884) was taken out from the -80℃ ultra-low temperature freezer and activated on a PDA plate, and cultured in a 28℃ incubator for 3 days. A 6cm long toothpick was sterilized and then evenly spread on the PDA medium in a 9cm culture dish. The sheath blight fungus was transferred to the medium and cultured for 4 days. Then the toothpick was placed 1cm below the third leaf sheath of the tillering stage rice (overexpression lines OsPLS3-1, OsPLS3-2, homozygous knockout mutant lines ospls3-1, ospls3-2) using tweezers, and then the leaf sheath was returned to normal, and the disease developed for 5 to 7 days, and the development of the disease was observed and recorded. The results of the inoculation showed that the lesion length of the OsPLS3 overexpression lines (OsPLS3-1, OsPLS3-2) was significantly shorter than that of the wild type ZH11, and thus the resistance of the OsPLS3 overexpression rice lines to sheath blight was significantly improved Figure 6), the disease resistance of homozygous knockout strain is lower than that of wild type.

[0104] V. Transgenic rice strain in vitro leaf inoculation verifies the resistance to rice sheath blight fungus:

[0105] The rice sheath blight fungus GD118 is activated, and the tillering stage of each transgenic strain (overexpression strain OsPLS3-1, OsPLS3-2, homozygous knockout mutant strain ospls3-1, ospls3-2) is also leaf. The middle part of the leaf is cut to about 10 cm, the wound site of the leaf is wounded using a gun head, and then a 5 mm mycelium block is placed at the wound site of the leaf, and the wound site is sealed with a plastic film. After being cultured in a 28℃ incubator for about 33 days, the disease condition is observed and photographed, and the disease spot area is counted using ImageJ. The results of the inoculation show that the disease spot area of the OsPLS3 overexpression strain OsPLS3-1 and OsPLS3-2 is significantly smaller than that of the wild type ZH11, and the disease spot area of the homozygous knockout mutant strain ospls3-1 and ospls3-2 is significantly larger than that of the wild type ZH11. Therefore, OsPLS3 positively regulates the resistance of rice to sheath blight ( Figure 7 ).

[0106] Experiments prove that overexpression of the rice adiponectin receptor gene OsPLS3 in rice in the present application can improve the resistance of rice to sheath blight, and knockout can make the rice more susceptible to sheath blight fungus. In the sheath blight fungus inoculation experiment on living plants, the OsPLS3 overexpression rice has a shorter disease spot length after being inoculated with sheath blight fungus than the wild type. After the sheath blight fungus is inoculated on the in vitro leaf of rice, the OsPLS3 overexpression rice has a smaller disease spot area than the wild type after the sheath blight fungus is inoculated on the leaf, and the ospls3 mutant has a larger disease spot. Therefore, OsPLS3 positively regulates the resistance of rice to sheath blight. The rice adiponectin receptor gene OsPLS3 in the present application has important production value for breeding rice resistant to sheath blight.

[0107] It should be noted that when numerical ranges are involved in the present application, both endpoints of each numerical range and any number between the two endpoints can be selected. Since the same steps and examples are used, the preferred embodiments are described in the present application to prevent redundancy. Although the preferred embodiments of the present application have been described, those skilled in the art can make additional changes and modifications to the embodiments once they know the basic creative concept. Therefore, the appended claims are intended to include the preferred embodiments and all changes and modifications falling within the scope of the present application.

[0108] Obviously, many modifications and variations of the present application are possible in light of the above teachings. It is, therefore, to be understood that within the scope of the appended claims and their equivalents, the application can be practiced otherwise than as specifically described.

Claims

1. The application of the rice adiponectin receptor OsPLS3 protein or its encoding gene shown in SEQ ID NO.3 in regulating plant disease resistance; wherein the plant is rice, and the disease resistance is resistance to sheath blight.

2. The application according to claim 1, characterized in that, The nucleotide sequence of the encoding gene is shown in SEQ ID NO.

1.

3. The application according to claim 1, characterized in that, The cDNA sequence of the encoding gene is shown in SEQ ID NO.

2.

4. The application of the rice adiponectin receptor OsPLS3 protein shown in SEQ ID NO.3 in the improvement of rice germplasm resources; the method of improving rice germplasm resources is to overexpress the encoding gene of the rice adiponectin receptor OsPLS3 protein in rice to enhance resistance to rice sheath blight; the encoding gene is shown in SEQ ID NO.1 or SEQ ID NO.

2.

5. A method for cultivating transgenic plants with enhanced disease resistance, comprising transferring the encoding gene described in SEQ ID NO.1 or SEQ ID NO.2 into a plant, and screening to obtain transgenic plants that overexpress the encoding gene, i.e., transgenic plants with enhanced disease resistance; wherein the plant is rice, and the disease resistance is resistance to rice sheath blight.

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