KR102478476B1 - LOC_Os09g04310 gene for controlling disease resistance of plant and uses thereof - Google Patents

Abstract

The present invention relates to the LOC_ Os09g04310 gene that regulates the disease resistance of plants and its use, and is expected to be useful for developing new plants resistant to white leaf blight by regulating the expression of the LOC_ Os09g04310 gene of the present invention. do.

Description

LOC_Os09g04310 gene for controlling disease resistance of plants and its uses {LOC_Os09g04310 gene for controlling disease resistance of plant and uses thereof}

The present invention relates to a LOC_ Os09g04310 gene that regulates plant disease resistance and uses thereof.

White leaf blight ( Xanthomonas oryzae pv. oryzae , Xoo) is a gram-negative bacterium, which is a major pathogen of white leaf blight in rice and is known as a major cause of bacterial diseases in rice worldwide. White leaf blight is infected through wounds or drainage tissues of host plants, proliferates through covering tissues, and moves through the vascular system. Correlation of diversity, distribution, phylogeny, and infectivity of white leaf blight obtained through molecular marker research helped to better understand the phylogenetic information of white leaf blight, showed that the shape of

The strains of the Korean type white blight showed phylogenetic differences of 64% and 58% from the Philippine type and the Japanese type, respectively. It is expected. The Korean K1 strain is on the decline due to the spread of rice cultivars that have the bacterial blight resistance genes Xa1 and Xa3 , but the K2 and K3 strains continue to proliferate in Korea. K3a, a new pathogen, shows the same infection pattern as K3, which caused severe white leaf blight damage in 2003.

Functional genetic analysis has proven essential to verify the biological significance of validated genes in examining non-specifically expressed protein genes in plants. Although several genes referred to as Xa genes exist, non-specific expressed proteins that are considered to be involved in the plant immune network through oligonucleotide microarray analysis of rice and white leaf blight are not known.

In order to identify important genes related to resistance to the Korean strain K2 of Xoo , the present inventors performed genome microarray analysis of japonica rice cultivars 'Dongjin' and 'Jinbaek' to confirm overexpression of several genes, among which LOC_ The Os09g04310 gene was an uncharacterized expressed protein. Therefore, the present invention was carried out to verify the function of the expressed protein in response to Xoo infection in rice.

On the other hand, Korean Patent Registration No. 1636154 discloses 'a methyltransferase resistant to white leaf blight and a transformed plant using the same', and Korean Patent Publication No. 2013-0055766 discloses 'a rice through gene expression suppression using a PSK08 gene derived from rice'. A method for imparting white leaf blight resistance is disclosed, but the LOC_Os09g04310 gene and its use for regulating the disease resistance of plants of the present invention are not described.

The present invention was derived from the above request, and the present inventors prepared a rice transformant overexpressing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 present in rice chromosome 9, and then white leaf blight pathogen ( Xanthomonas oryzae pv. oryzae strain K2) was inoculated and the degree of plant damage caused by the white leaf blight was observed. As a result, the rice transformants overexpressing the LOC_ Os09g04310 gene were superior to the wild-type Dongjin rice (susceptibility to white leaf blight). It was confirmed that the length of the lesion was reduced, and through this, it was confirmed that the LOC_ Os09g04310 gene could increase the resistance of plants to white leaf blight, thereby completing the present invention.

In order to solve the above problems, the present invention is a plant disease resistance of plants comprising the step of regulating the expression of the LOC_ Os09g04310 gene by transforming plant cells with a recombinant vector containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 provides a way to regulate

In addition, the present invention comprises the steps of transforming a plant cell with a recombinant vector containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1; and regenerating plants from the transformed plant cells.

In addition, the present invention provides a transgenic plant with controlled plant disease resistance prepared by the above production method and a transformed seed thereof.

In addition, the present invention provides a composition for regulating plant disease resistance of plants containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 as an active ingredient.

Since the transgenic plants overexpressing the LOC_ Os09g04310 gene of the present invention have increased resistance to white leaf blight, the productivity of crops can be improved by developing transgenic plants with controlled resistance to white leaf blight by controlling the expression of the LOC_ Os09g04310 gene. hope to be able to

1 is a schematic diagram of a recombinant vector containing a LOC_Os09g04310 gene composed of the nucleotide sequence of SEQ ID NO: 1.
2 is a result of comparing the nucleotide sequences of the LOC_ Os09g04310 gene (REF-NB) in the rice standard genome (Nipponbare cultivar) and the LOC_ Os09g04310 gene cloned in the present invention.
Figure 3 shows the rice transformants overexpressing the LOC_ Os09g04310 gene by tissue (root, stem; stem; leaf sheath; leaf, leaf; and flag leaf) and growth period (germination, germination stage; This is the PCR result confirming the expression level of the LOC_ Os09g04310 gene according to three-leaf stage, 3-leaf stage; maximum tillering stage; panicle initiation stage).
4 is a PCR result confirming the copy number of the inserted gene in the rice transformant overexpressing the LOC_ Os09g04310 gene. NC (Negative control): Dongjinbyeop, PC (Positive control): mock vector (pCAMBIA1300 vector without LOC_ Os09g04310 gene inserted).
Figure 5 is a white leaf blight susceptible variety, wild type Dongjinbyeo (DJ), white leaf blight resistant variety, Jinbaekbyeo (JB), and LOC_ Os09g04310 gene overexpressing rice transformants ( LOC_ Os09g04310 OX-1, 2 and 3). oryzae pv. oryzae strain K2) was inoculated and the lesion length of each plant was measured 8 days, 12 days and 16 days later.
6 is a white leaf blight susceptible variety, wild type Dongjin rice (Dongjin), a white leaf blight resistant variety, Jinbaek rice ( Jinbaek ), and a white leaf blotch pathogen ( X This is a photograph of the degree of damage to white leaf blight in each plant after inoculation with oryzae pv. oryzae strain K2).

In order to achieve the object of the present invention, the present invention is a plant plant comprising the step of regulating the expression of the LOC_ Os09g04310 gene by transforming plant cells with a recombinant vector containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 A method for regulating disease resistance is provided.

The method of regulating plant disease resistance of plants according to the present invention may be to overexpress the LOC_ Os09g04310 gene in plant cells to increase plant disease resistance, but is not limited thereto.

In the method according to an embodiment of the present invention, the plant disease may be a wide range of plant diseases such as white leaf blight, blast disease, striped leaf blight, leaf sheath disease, black streak disease, spot blotch disease, soft rot, wilt disease, warp disease, and ulcer disease, Preferably, it may be white leaf blight caused by a white leaf blight pathogen ( Xanthomonas oryzae pv. oryzae strain K2), but is not limited thereto.

In addition, in the method for controlling plant disease resistance of a plant according to an embodiment of the present invention, the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 is a reference sequence (http://rice.plantbiology.msu.edu/ cgi-bin/sequence_display.cgi?orf=LOC_Os09g04310) and a sequence having 100% homology with the coding sequence (CDS) (see FIG. 2).

The scope of the LOC_ Os09g04310 gene of the present invention includes genomic DNA, cDNA and synthetic DNA of the LOC_ Os09g04310 gene. Preferably, the LOC_ Os09g04310 gene of the present invention may include the nucleotide sequence represented by SEQ ID NO: 1. In addition, homologues of the above nucleotide sequences are included within the scope of the present invention. Specifically, the gene includes a base sequence having 70% or more, more preferably 80% or more, more preferably 90% or more, and most preferably 95% or more sequence homology with the nucleotide sequence of SEQ ID NO: 1, respectively. can do. The "% sequence homology" for polynucleotides is determined by comparing two optimally aligned sequences, wherein a portion of the polynucleotide sequence in the region of comparison is a reference sequence (not containing additions or deletions) to the optimal alignment of the two sequences. may include additions or deletions (i.e., gaps) compared to

As used herein, the term "recombinant" refers to a cell that replicates, expresses a heterologous nucleic acid, or expresses a peptide, a protein encoded by a heterologous peptide, or a heterologous nucleic acid. Recombinant cells can express genes or gene segments not found in the cell's native form, either in sense or antisense form. A recombinant cell may also express a gene found in the cell in its natural state, but the gene has been reintroduced into the cell by artificial means as a modified one.

In this specification, the term "vector" is used to refer to DNA fragment(s) or nucleic acid molecules that are delivered into cells. Vectors replicate DNA and can reproduce independently in host cells. The term “delivery vehicle” is often used interchangeably with “vector”.

Vectors of the invention may typically be constructed as vectors for expression or cloning. In addition, the vector of the present invention can be constructed using a prokaryotic or eukaryotic cell as a host. For example, when the vector of the present invention is an expression vector and a prokaryotic cell is used as a host, a strong promoter capable of promoting transcription (eg, pLλ promoter, trp promoter, lac promoter, T7 promoter, tac promoter, etc.) ), a ribosome binding site for initiation of translation, and a transcription/translation termination sequence. When Escherichia coli is used as a host cell, promoter and operator sites of the E. coli tryptophan biosynthetic pathway and leftward promoter (pLλ promoter) of phage λ may be used as control sites.

In the recombinant vector of the present invention, the promoter may be promoters suitable for transformation, preferably CaMV 35S promoter, actin promoter, ubiquitin promoter, pEMU promoter, MAS promoter or histone promoter, preferably CaMV 35S promoter. However, it is not limited thereto.

As used herein, the term "promoter" refers to a region of DNA upstream from a structural gene and refers to a DNA molecule to which RNA polymerase binds to initiate transcription. A "plant promoter" is a promoter capable of initiating transcription in a plant cell. A “constitutive promoter” is a promoter that is active under most environmental conditions and states of development or cell differentiation. Constitutive promoters may be preferred in the present invention because selection of transformants may be made by various tissues at various stages. Thus, constitutive promoters do not limit selection possibilities.

The recombinant vector of the present invention can be constructed by methods well known to those skilled in the art. The method includes in vitro recombinant DNA technology, DNA synthesis technology, in vivo recombinant technology, and the like. The DNA sequence can be effectively linked to a suitable promoter in an expression vector to direct mRNA synthesis. In addition, the vector may include a ribosome binding site and a transcription terminator as a translation initiation site.

A preferred example of the recombinant vector of the present invention is a Ti-plasmid vector capable of transferring a part of itself, the so-called T-region, into plant cells when present in a suitable host such as Agrobacterium tumefaciens . Another type of Ti-plasmid vector (see EP 0 116 718 B1) is currently used to transfer hybrid DNA sequences into plant cells, or protoplasts, from which new plants can be produced that properly integrate the hybrid DNA into the plant's genome. there is. A particularly preferred form of the Ti-plasmid vector is the so-called binary vector as claimed in EP 0 120 516 B1 and US Pat. No. 4,940,838. Other suitable vectors that can be used to introduce DNA according to the present invention into plant hosts include viral vectors, such as those that can be derived from double-stranded plant viruses (eg, CaMV) and single-stranded viruses, gemini viruses, and the like. For example, it may be selected from incomplete plant viral vectors. The use of such vectors can be particularly advantageous when properly transforming a plant host is difficult.

Recombinant expression vectors may preferably contain one or more selectable markers. The marker is a nucleic acid sequence having a characteristic that can be selected by a conventional chemical method, and includes all genes capable of distinguishing transformed cells from non-transformed cells. The marker gene may be an antibiotics resistance gene, but is not limited thereto.

In the recombinant vector of the present invention, it is possible to use a conventional terminator, for example, a nopaline synthase (NOS) terminator, a rice α-amylase (RAmy1 A) terminator, a phaseoline terminator, Agrobacterium tumefaciens ( Agrobacterium tumefaciens ) of octopine synthase (Octopine synthase) gene terminator and the like, but is not limited thereto. Regarding the need for terminators, it is generally known that such regions increase the certainty and efficiency of transcription in plant cells. Therefore, the use of terminators is highly desirable in the context of the present invention.

Any host cell known in the art can be used as a host cell capable of stably and continuously cloning and expressing the vector of the present invention, and examples of prokaryotic cells include E. coli JM109, E. coli BL21, and E. coli. Bacillus genus strains such as RR1, E. coli LE392, E. coli B, E. coli X 1776, E. coli W3110, Bacillus subtilis , Bacillus thuringiensis , and Salmonella typhimurium Enterobacteriaceae and strains such as Salmonella typhimurium , Serratia marcescens and various Pseudomonas species.

When the vector of the present invention is transformed into a eukaryotic cell, as a host cell, yeast ( Saccharomyce cerevisiae ), insect cell, human cell (eg, CHO cell line (Chinese hamster ovary), W138, BHK, COS-7, 293, HepG2 , 3T3, RIN and MDCK cell lines) and plant cells, etc. may be used. The host cell is preferably a plant cell.

Methods for delivering the vector of the present invention into a host cell, when the host cell is a prokaryotic cell, include the CaCl ₂ method, the Hanhan method (Hanahan, D., 1983 J. Mol. Biol. 166, 557-580), and electroporation. It can be implemented by methods and the like. In addition, when the host cell is a eukaryotic cell, the vector can be injected into the host cell by microinjection, calcium phosphate precipitation, electroporation, liposome-mediated transfection, DEAE-dextran treatment, and gene bombardment. can

The present invention also includes the steps of transforming plant cells with a recombinant vector containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1; and

It provides a method for producing a transgenic plant with controlled plant disease resistance, comprising: regenerating a plant from the transformed plant cell.

In the method for producing a transgenic plant with controlled plant disease resistance of the present invention, the range of the LOC_ Os09g04310 gene is the same as described above.

In the production method according to one embodiment of the present invention, the method for producing a transgenic plant with controlled plant disease resistance may be to increase the resistance of the plant to plant disease by overexpressing the LOC_ Os09g04310 gene in plant cells. Not limited to this.

In the manufacturing method according to the present invention, the plant disease may preferably be white leaf blight caused by a white leaf blight pathogen ( Xanthomonas oryzae pv. oryzae strain K2), but is not limited thereto.

Plant transformation refers to any method of transferring DNA into a plant. Such transformation methods need not necessarily have a period of regeneration and/or tissue culture. Transformation of plant species is now common for plant species including both dicotyledonous as well as monocotyledonous plants. In principle, any transformation method can be used to introduce the hybrid DNA according to the present invention into suitable progenitor cells. The method is the calcium/polyethylene glycol method for protoplasts (Krens, F.A. et al., 1982, Nature 296, 72-74; Negrutiu I. et al., 1987, Plant Mol. Biol. 8, 363-373), Electroporation (Shillito R.D. et al., 1985, Bio/Technol. 3, 1099-1102), microinjection into plant elements (Crossway A. et al., 1986, Mol. Gen. Genet. 202, 179-185 ), Agrobacterium tumefacies by particle bombardment of various plant elements (DNA or RNA-coated) (Klein T.M. et al., 1987, Nature 327, 70), infiltration of plants or transformation of mature pollen or microspores It can be appropriately selected from infections by viruses in (incomplete) ens-mediated gene transfer (EP 0 301 316) and the like. Preferred methods according to the present invention include Agrobacterium mediated DNA delivery. Particularly preferred is the use of so-called binary vector techniques as described in EP A 120 516 and US Pat. No. 4,940,838.

In addition, as a method for regenerating a transgenic plant from the transformed plant cell, any method known in the art may be used.

The present invention also provides a transgenic plant with controlled plant disease resistance prepared by the above production method and a transformed seed thereof.

The transgenic plant of the present invention may be one in which the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 is overexpressed in plant cells to increase resistance to plant diseases compared to non-transformed plants, but is not limited thereto.

In addition, the plant disease may preferably be white leaf blight caused by a white leaf blight pathogen ( Xanthomonas oryzae pv. oryzae strain K2), but is not limited thereto.

The plants are rice, barley, wheat, rye, corn, sugarcane, oats, onion, or Arabidopsis monocotyledonous plants, potatoes, eggplants, tobacco, peppers, tomatoes, burdock, crown daisy, lettuce, bellflower, spinach, chard, sweet potato , Dicotyledons such as carrots, parsley, Chinese cabbage, cabbage, mustard radish, watermelon, melon, cucumber, pumpkin, gourd, strawberry, soybean, mung bean, kidney bean, pea, etc., preferably a monocotyledonous plant, more preferably It may be rice, but is not limited thereto.

The present invention also provides a composition for regulating plant disease resistance containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 as an active ingredient. The composition of the present invention contains, as an active ingredient, the LOC_ Os09g04310 gene capable of controlling the resistance to white leaf blight caused by the plant's white leaf blight pathogen ( Xanthomonas oryzae pv. oryzae strain K2), and the expression of the LOC_ Os09g04310 gene is increased. It can increase the resistance of plants to white leaf blight.

Hereinafter, the present invention will be described in detail by examples. However, the following examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following examples.

Materials and Methods

1. LOC_ Os09g04310 gene isolation

Total RNA was extracted with RNAiso Plus extraction reagent (Takara Bio Inc., Japan) using young leaves of 'Jinbaek', a rice variety that is resistant to white leaf blight, and about 1 ug of the extracted RNA was used as a template for Superscript III First-Strand cDNA Synthesis cDNA was synthesized with Kit (Invitrogen, USA), and the primer set shown in Table 1 was used for PCR amplification of the gene (Table 1). The PCR process was pre-denaturation at 95° C. for 5 minutes; The process of denaturation at 95°C for 30 seconds, annealing at 55°C for 30 seconds, and extension at 72°C for 2 minutes was repeated a total of 35 times; Final elongation at 72°C for 10 minutes; To confirm the PCR product, electrophoresis was performed using a 1% (w/v) agarose gel containing ethidium bromide, and the PCR product of the expected size was extracted from the agarose gel and purified using a Gel purification kit (Bioneer , USA) was used for purification. The purified PCR product was subcloned into a pGEMT-easy vector (Promega, USA) and sequenced by Macrogen (Macrogen, Korea). Base sequence homology was confirmed through multiple base sequence alignment by hierarchical clustering analysis.

Primer information used in the present invention Gene ID Seq(5'→3') (SEQ ID NO) restriction enzyme LOC_ Os09g04310 F: ggtaccATGGCAGCCAGAACAAGCGCTA (2) Kpn I R: tctagaTTAGCAATTTGCTTCACCGGTAGG (3) Xba I

Lower case: Restriction enzyme recognition site

2. Construction of overexpressing recombinant vector

The full-length cDNA of the LOC_ Os09g04310 gene was inserted between the restriction enzymes Kpn I and Xba I in the pCAMBIA1300 vector (Takara, Japan) using the T4 ligase kit (Promega, USA). The hptII ( hygromycin phosphotransferase II) gene was used as a plant selection marker, and the 35S promoter was used to induce overexpression of the inserted gene (FIG. 1).

3. Production of rice transformants

The completed vector was transformed into EHA105, an Agrobacterium tumefaciens strain, and the transformed Agrobacterium was selected by culturing in a 1.5% AB medium containing kanamycin (50 mg/L) at 28°C in the dark. . Transformed Agrobacterium was suspended in AAM medium, adjusted to a final concentration of OD ₆₀₀ of 0.01, and then used in the experiment. Rice seed-derived callus cultured for 10 days was infected with Agrobacterium suspension for 3 minutes, dried, and then cultured in 2N6-AS medium containing 0.4% gelrite. After co-culture, callus was washed with distilled water containing carbenicillin (500 mg/L) to remove Agrobacterium. The washed callus was cultured in 2N6-CP medium containing hygromycin (50 mg/L) and carbenicillin (400 mg/L) under continuous light conditions at 32°C for 2 weeks. Thereafter, the proliferated callus was transferred to MSR-CP medium to induce stem and root elongation, and continuous subculture was performed. The regenerated plants from the callus were transferred to soil and grown in a greenhouse for 2 weeks. The composition of the medium used for tissue culture and transformation is as shown in Table 2 of Korean Registered Patent No. 2103189.

4. Genomic DNA extraction

Total genomic DNA was extracted using the TissueLyser II system (QIAGEN, UK) and CTAB (Cetyl Trimethyl Ammonium Bromide) using young leaves 2 weeks after transplanting. After DNA extraction, the purity and concentration of DNA was measured using a spectrophotometer (NanoDrop ^TM One, Thermo Fisher Scientific, USA), and the extracted DNA was diluted with sterile water and stored at -20 °C before use.

5. Copy number analysis through TaqMan qPCR

TaqMan assay (Applied Biosystems, USA) was performed to confirm the copy number of the gene inserted into the T ₀ transformant. 10 ng of DNA was mixed with FAM dye-labeled MGB probe for target gene sequence detection, VIC dye-labeled TAMRA probe for reference gene sequence detection, TaqMan Genotyping Master Mix containing AmpliTaq Gold DNA polymerase and dNTPs, All qPCR reactions were performed in triplicate using ABI 7900HT equipment (Alpplied Biosystems, USA), followed by denaturation at 95 °C for 10 minutes, followed by 15 seconds at 95 °C and 60 seconds at 60 °C. For data analysis, CopyCaller Software v 2.0 (Applied Biosystems, USA) was used.

6. LOC_ Os09g04310 Expression analysis of

To analyze the expression of the LOC_ Os09g04310 gene by tissue and growth period of transformants, RNA was analyzed in various tissues (root, stem, leaf sheath, leaf, branch) and growth period (germination, trifoliate, maximal division, ear formation). extracted.

7. Verification of resistance to white leaf blight

LOC_ Os09g04310 gene After sowing the overexpressing rice transformants, wild-type Dongjinbyeo (white leaf blight susceptible) and Jinbaek rice (white leaf blight resistance), 7-week-old plants were inoculated with the white leaf blotch pathogen ( X. oryzae pv. oryzae strain K2, 10 ⁸ CFU/㎖) After 8 days, 12 days, and 16 days, the length of the leaf tissue was measured to determine the degree of infectivity. Each was repeated 5 times.

Example 1. Cloned LOC_ Os09g04310 Characteristics of genes

As a result of sequencing after cloning the LOC_ Os09g04310 gene from Jinbaek rice, it was confirmed that it was a non-annotated gene encoding a non-specific expressed protein. In addition, the LOC_ Os09g04310 gene cloned in the present invention is located on chromosome 9 of rice, and its molecular weight and isoelectric point were predicted to be 34.788 kDa and 8.9587, respectively, and have an open reading frame (ORF) of 951 bp encoding 317 amino acid residues. Confirmed.

In addition, the nucleotide sequence of LOC_ Os09g04310 in the rice standard genome (Nipponbare cultivar) (http://rice.plantbiology.msu.edu/cgi-bin/sequence_display.cgi?orf=LOC_Os09g04310) and the LOC_ Os09g04310 gene cloned from Jinbaek rice Comparing the nucleotide sequences of , it was confirmed that they showed 100% homology (FIG. 2).

Example 2. Cloned LOC_ Os09g04310 Spatio-temporal specific expression of genes

As a result of analyzing the expression level of the LOC_ Os09g04310 gene according to tissue and growth period of rice transgenic plants, leaf sheath, leaf, and flag leaf compared to root and stem. It was confirmed that the expression of the LOC_ Os09g04310 gene was relatively high in , LOC_ Os09g04310 depending on the growth period (germination stage; 3-leaf stage; maximum tillering stage; panicle initiation stage). It was confirmed that the expression of the gene increased (FIG. 3).

Example 3. Development of overexpressing rice transformants

After transforming 'Dongjin', a japonica rice cultivar, with an overexpression recombinant vector into which the full-length cDNA of the LOC_ Os09g04310 gene was inserted, the T ₀ transformant was used to target the inserted gene and HPT marker to confirm whether the recombinant vector was inserted. PCR analysis was performed to confirm successful introduction of the LOC_ Os09g04310 gene (data not shown). In addition, TaqMan qPCR was performed to confirm the copy number of the inserted gene using the gDNA extracted from the T ₀ transformant, and LOC_ Os09g04310 -1, 2 and 3 strains having a single copy number were selected. (FIG. 4).

Example 4. Resistance assay for white leaf blight of rice transformants

After inoculating rice transformants with LOC_ Os09g04310 gene overexpression (LOC_ Os09g04310 -1, 2 and 3) with white leaf blight pathogen K2, the lesion length of each plant was measured on the 8th, 12th and 16th days The effect of LOC_ Os09g04310 gene overexpression on white leaf blight resistance was analyzed.

As a result, on the 8th day of inoculation, it was confirmed that the lesion length of all transformants was similar to that of wild-type Dongjinbyeo (susceptibility to white leaf blight). However, on the 12th day of inoculation, the lesion length of wild-type Dongjinbyeo was confirmed to be about 13 cm, and the lesion length of the transformant was about 6.5 to 8.5 cm. On the 16th day of inoculation, the lesion length of wild-type Dongjinbyeo was about 19 cm, The length of the lesion on the sieve was confirmed to be about 8.5 to 9.9 cm, and it was confirmed that the degree of damage to white leaf blight was very serious in wild-type Dongjinbyeo compared to the LOC_ Os09g04310 gene overexpressing rice transformants. On the other hand, Jinbaek rice, a variety resistant to white leaf blight, had the shortest lesion length compared to other experimental groups (FIGS. 5 and 6).

Through the above results, it can be seen that resistance to white leaf blight is increased when the LOC_ Os09g04310 gene of the present invention is overexpressed in plants, and it is thought that the resistance to white leaf blight of plants can be controlled by controlling the expression of the LOC_ Os09g04310 gene It became.

<110> Chungbuk National University Industry-Academic Cooperation Foundation <120> LOC_Os09g04310 gene for controlling disease resistance of plant and uses it <130> PN20176 <160> 3 <170> KoPatentIn 3.0 <210> 1 <211> 951 <212> DNA <213> Oryza sativa <400> 1 atggcagcca gaacaagcgc taacttctca tcatgcttta ccttcctgaa ggaagcccta 60 gtcctcccaa ctcagaaccc caagctcttc acacctgtct tcctcctgat cgcgcttcct 120 agcttccttg tcctctctac caatgtccta ttcgtccagc cactctccat ggacatggcc 180 gaacttgcaa tcaagctcca gaccaccgac ccttccagcg ccgagtaccg catgatcctg 240 gaggagctga aacatgacgt tacacagctc atcctcgtcg tcgtcgccgt cgagctcgtc 300 gctttggtgc ttggcttcgt caatcagtgc gtcggcttct tcgcggcctc ctccacctac 360 tccggcgacc gctactcgct cccggagctc ctccgcaagg cgatgaaggg gaatctcaag 420 ggtcctctca taaccatcgc catggtcacc gtactccggg tcacatacat ggcgctcctc 480 ggtgttctaa tctattccgt catgcaagtg caacggcact acttgatcaa ggtgctctcc 540 gtgcaagtcc tcctcttcgt cctctgtttc ctcgccttcc tctacttcaa cgtcgttggc 600 atggtgagcg tcgccgtgtc ggtgggcgac acggagcgcc gcgggatccg ggcgctccgg 660 caggcgtggc ggctgatgac gcgggtgagg aggaaggagg gcctcgtgct ggtggtggcc 720 atctgcctct tgtcgatagc tgtcagcccg gtgaacctgg tcgctgctgc gtacacgaag 780 aacatggtac tggggctatg cctcttggtc gtctatgctc tgctgtctgg tgcagagcag 840 ctgttctact ttgcagcagc cacggtgtac tactgccaag caatggacag caagggagag 900 gccatggatt atgcatacgc caagatccct accggtgaag caaattgcta a 951 <210> 2 <211> 28 <212> DNA <213> artificial sequence <220> <223> primer <400> 2 ggtaccatgg cagccagaac aagcgcta 28 <210> 3 <211> 30 <212> DNA <213> artificial sequence <220> <223> primer <400> 3 tctagattag caatttgctt caccggtagg 30

Claims (9)

A method for increasing bacterial leaf blight disease resistance of a plant comprising the step of over-expressing the expression of the LOC_ Os09g04310 gene by transforming plant cells with a recombinant vector containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 . delete delete Transforming plant cells with a recombinant vector containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 to overexpress the LOC_ Os09g04310 gene in the transformed plant cells; and
A method for producing a transgenic plant with increased resistance to bacterial leaf blight disease, comprising: regenerating a plant from the transformed plant cell. delete A transgenic plant with increased resistance to bacterial leaf blight disease prepared by the method of claim 4. delete Transformed seeds of plants according to claim 6. A composition for increasing bacterial leaf blight disease resistance of a plant containing the LOC_ Os09g04310 gene consisting of the nucleotide sequence of SEQ ID NO: 1 as an active ingredient.

Images