CN117802151B - Application of rice root-knot nematode disease gene OsThil in aspect of regulating and controlling resistance of rice to root-knot nematodes - Google Patents

Abstract

The invention discloses an application of a rice root-knot nematode disease gene OsThil in regulating and controlling the resistance of rice to root-knot nematodes, and belongs to the technical field of biology. The invention utilizes CRISPR/Cas9 gene editing technology and gene over-expression technology, through agrobacterium-mediated mutant system, site-directed mutation and over-expression of infectious disease genes OsThil are respectively carried out in rice, and rice explants are transformed to obtain rice mutant plants with OsThil gene function deletion and OsThil gene expression increase respectively, wherein the OsThil gene function deletion mutant can obviously enhance the resistance of the rice to root-knot nematodes. The OsThil gene disclosed by the invention is utilized to construct the gene deletion mutant, which is beneficial to improving the capability of resisting pathogen infection of crop varieties and has important significance for expanding the planting range of crops and improving the crop yield.

Description

Application of rice root-knot nematode susceptibility gene OsThil in regulating rice resistance to root-knot nematodes

Technical Field

The invention relates to the field of biotechnology, and in particular to application of a rice root-knot nematode susceptibility gene OsThil in regulating the resistance of rice to the root-knot nematode.

Background technique

Rice (Oryza sativa L.) is one of the main food crops of the world's population. Rice root-knot nematode disease is currently the most serious nematode disease on rice, and the rice yield loss caused by rice root-knot nematode disease is as high as 20% to 80% each year. Production practice shows that exploring the genetic resources of rice susceptible to rice root-knot nematodes, using gene editing technology to destroy the susceptible genes in susceptible rice varieties, and then cultivating disease-resistant rice varieties is an effective, economical, environmentally friendly and lasting strategy for preventing and controlling rice root-knot nematodes.

Rice root-knot nematode susceptibility genes are controlled by quantitative traits, and currently few root-knot nematode susceptibility genes have been located in rice. Only a few quantitative trait loci for resistance to rice root-knot nematodes have been discovered in rice. The cloning of rice root-knot nematode resistance genes has promoted the development of molecular improvement of rice resistance to rice root-knot nematodes, but the resistance spectrum of rice resistance genes that have been located or cloned is not broad. Rice root-knot nematodes can overcome rice resistance genes and then infect rice. Therefore, destroying rice root-knot nematode susceptibility genes has become a hot topic in recent years, providing a new solution for cultivating rice root-knot nematode resistance genetic breeding.

Genome-wide association analysis is a statistical analysis of the correlation between genotype and phenotype at the genome-wide level using molecular genetic markers combined with phenotypic traits, in order to discover and locate genes and variants that affect complex traits. With the rapid development of high-throughput sequencing technology, it has become possible to use tens of thousands of single nucleotide polymorphisms in the rice genome as molecular genetic markers, which has greatly promoted the application of genome-wide association analysis in the location and cloning of rice disease susceptibility genes.

Recently, CRISPR/Cas9 gene editing technology has been widely applied to rice disease-susceptibility genes, enabling the breeding of broad-spectrum disease-resistant rice varieties. For example, Xu et al. used CRISPR/Cas9 technology to precisely edit the 3’UTR region of the disease-susceptibility genes OsRNG1 and OsRNG3 of the rice variety Nipponbare, and obtained four mutant strains with significant resistance to rice blast, and the other agronomic traits of the mutant strains were consistent with those of the parents. However, there are few reports on methods for mutating disease-susceptibility genes to improve rice resistance to root-knot nematodes.

Summary of the invention

The present invention aims to provide an application of a rice root-knot nematode susceptibility gene OsThil in regulating the resistance of rice to root-knot nematodes, so as to solve the problems existing in the above-mentioned prior art. The deletion of the rice root-knot nematode susceptibility gene OsThil can significantly improve the resistance of rice to root-knot nematodes.

To achieve the above object, the present invention provides the following solutions:

The present invention provides the use of the rice root-knot nematode susceptibility gene OsThil in the following (1) or (2):

(1) Application in regulating rice resistance to root-knot nematodes;

(2) Application in breeding transgenic rice with enhanced susceptibility or resistance to rice root-knot nematodes;

Wherein, the nucleotide sequence of OsThil is shown as SEQ ID NO.1.

The present invention also provides the use of the protein expressed by the rice root-knot nematode susceptibility gene OsThil in the following (1) or (2):

(1) Application in regulating rice resistance to root-knot nematodes;

(2) Application in breeding transgenic rice with enhanced susceptibility or resistance to rice root-knot nematodes.

The present invention also provides the use of a recombinant vector comprising the rice root-knot nematode susceptibility gene OsThil in the following (1) or (2):

(1) Application in regulating rice resistance to root-knot nematodes;

(2) Application in breeding transgenic rice with enhanced susceptibility or resistance to rice root-knot nematodes.

The present invention also provides the use of a host cell comprising the recombinant vector in the following (1) or (2):

(1) Application in regulating rice resistance to root-knot nematodes;

(2) Application in breeding transgenic rice with enhanced susceptibility or resistance to rice root-knot nematodes.

Preferably, the resistance of rice to root-knot nematodes is enhanced by reducing or interfering with the expression of the rice root-knot nematode susceptibility gene OsThil or a fragment thereof, or knocking out the rice root-knot nematode susceptibility gene OsThil or a fragment thereof; overexpressing the rice root-knot nematode susceptibility gene OsThil or a fragment thereof enhances the susceptibility of rice to root-knot nematodes. The above-mentioned knockout fragment of the rice root-knot nematode susceptibility gene OsThil includes but is not limited to the sequence after knocking out positions 23-27 (CTTCG) or positions 24-25 (TT) of the sequence shown in SEQ ID NO.1.

The present invention also provides a method for cultivating transgenic rice with enhanced susceptibility to root-knot nematodes, comprising overexpressing a rice root-knot nematode susceptibility gene OsThil in rice to cultivate transgenic rice with enhanced susceptibility to root-knot nematodes; the nucleotide sequence of the rice root-knot nematode susceptibility gene OsThil is shown in SEQ ID NO.1.

The present invention also provides a method for cultivating transgenic rice with enhanced resistance to root-knot nematodes, comprising knocking out the rice root-knot nematode susceptibility gene OsThil or a fragment thereof in rice to cultivate transgenic rice with enhanced resistance to root-knot nematodes; the nucleotide sequence of the rice root-knot nematode susceptibility gene OsThil is shown in SEQ ID NO.1.

The present invention discloses the following technical effects:

The present invention uses CRISPR/Cas9 gene editing technology and overexpression technology to obtain Nipponbare rice OsThil gene mutant plants, and verifies a root-knot nematode susceptible gene OsThil gene in rice through indoor inoculation. The gene can mediate root-knot nematode infection of rice. The research results show that overexpression of OsThil gene significantly enhances the sensitivity of plants to nematodes, while knocking out OsThil gene can significantly improve the resistance of rice to root-knot nematodes, and will not affect the growth phenotype of rice plants. The cultivation method of transgenic rice constructed by CRISPR/Cas9 gene editing technology can realize the expansion of rice resistance resources, and can conveniently and efficiently apply fixed-point editing of rice susceptible gene sites to improve resistance to root-knot nematodes, and obtain new root-knot nematode resistance rice lines with OsThil gene function loss, which provides germplasm resources and creation methods for green prevention and control of rice root-knot nematodes and molecular breeding of resistant varieties.

BRIEF DESCRIPTION OF THE DRAWINGS

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

Figure 1 shows the OsThil gene structure and guide RNA (gRNA) target sequence;

Figure 2 is a schematic diagram of the T-DNA structure of the pYLCRISPR/Cas9-Pubi-H-OsThil-CR vector;

FIG3 is a comparison result of the target sequence mutation (a) and amino acid change (b) of the OsThil gene of the mutant Thil-CR;

Figure 4 is a schematic diagram of the T-DNA structure of the pRHV-OsThil-OE vector;

Figure 5 shows the results of positive screening of Thil-OE mutant plants by hygromycin; WT is a wild-type plant; Thil-OE is an overexpression mutant plant;

Figure 6 shows the phenotypic results of the deletion mutant Thil-CR and the overexpression mutant Thil-OE and Nipponbare plants; a: plant height comparison results; b: root length comparison results;

Figure 7 shows the statistical results of nematode infection and development after the deletion mutant strain Thil-CR, the overexpression mutant strain Thil-OE and the Nipponbare plants were inoculated with the root-knot nematodes of the Poaceae family, with the wild type Nipponbare as the control; a: statistical results of the number of root knots of each plant; b: statistical results of the number of root-knot nematodes infected in each plant; c: statistical results of the proportion of root-knot nematodes of the Poaceae family at different developmental stages; * indicates that the t-test test has significant differences (P<0.05); ** indicates that the t-test test has extremely significant differences (P<0.05).

Detailed ways

Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as commonly understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe methods and/or materials related to the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

The inventors previously located the OsThil gene within the 4.5 Mbp segment of the molecular marker on chromosome 7 of rice through whole genome association combined with transcriptome analysis, which is closely related to the resistance to root-knot nematodes. In order to further analyze the function of the gene in the interaction between rice and root-knot nematodes, the present invention uses CRISPR/Cas9 gene editing technology and overexpression technology to obtain Nipponbare rice OsThil gene mutant plants, and verifies a root-knot nematode susceptible gene OsThil gene in rice through indoor inoculation. The gene can mediate root-knot nematode infection of rice. Studies have shown that by overexpressing the OsThil gene, a rice mutant plant with increased OsThil gene expression is obtained, and the sensitivity of the plant to root-knot nematodes is significantly increased, indicating that the OsThil gene is a rice root-knot nematode susceptible gene and can significantly enhance the sensitivity of the plant to nematodes; and a CRISPR/Cas9 plant expression vector containing the OsThil gene sgRNA sequence is constructed, and the OsThil gene target site is mutated in rice through the Agrobacterium-mediated genetic transformation system to obtain a rice plant with a stable inheritance of OsThil gene function loss, and the resistance of the plant to root-knot nematodes is significantly improved. This indicates that knocking out the OsThil gene can significantly improve rice's resistance to root-knot nematodes without affecting the growth phenotype of rice plants.

In order to explain the above technical solution in more detail, a specific embodiment is further described below.

The gene sequences and primer sequences involved in the following examples are shown in Table 1 below.

Table 1 Gene and primer sequences

Example 1: Site-directed knockout of rice disease susceptibility gene OsThil based on CRISPR/Cas9 gene editing technology

1. Design of gRNA target sites

According to the sequence of the OsThil gene as shown in SEQ ID NO.1, a target site gRNA was designed on the exon of the OsThil gene, and its sequence was shown in SEQ ID NO.2 (as shown in FIG. 1 ).

2. Construction of plant expression vector pYLCRISPR/Cas9-Pubi-H-OsThil-CR

Using Overlapping PCR method:

The main purpose of the first round of PCR is to introduce gRNA into the downstream of OsU3 promoter and upstream of sgRNA sequence using pYLsgRNA-OsU3 as template. Reaction 1 is to introduce gRNA sequence into the downstream of OsU3 promoter after amplification using primer pair U-F/OsThil-U3T1 (sequence as shown in SEQ ID NO.13); Reaction 2 is to introduce gRNA sequence into the upstream of sgRNA sequence after amplification using primer pair gR-R/OsThil-gRT1 (sequence as shown in SEQ ID NO.14).

The amplification reaction system is shown in Table 2 below:

Table 2 First round amplification reaction system

The reaction program was: 95°C for 5 min; 95°C for 10 s, 58°C for 15 s, 72°C for 15 s, 25 cycles; 72°C for 10 min.

Subsequently, the first round of PCR products (products of reaction 1 and reaction 2) were used as templates, and the promoter, target site and sgRNA backbone were constructed into a complete expression cassette using primer pair Pps/Pgs. The sequence after amplification is shown in SEQ ID NO.15.

The amplification reaction system is shown in Table 3 below:

Table 3 Second round amplification reaction system

The reaction program was: 95°C for 5 min; 95°C for 10 s, 58°C for 15 s, 72°C for 20 s, 28 cycles; 72°C for 10 min.

Then, the gRNA expression cassette was cloned into pYLCRISPR/Cas9-Pubi-H to obtain the recombinant plant expression vector pYLCRI SPR/Cas9-Pubi-H-OsThil-CR, and its T-DNA structure diagram is shown in Figure 2.

The vectors pYLsgRNA-OsU3 and pYLCRISPR/Cas9-Pubi-H are derived from the Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, and have been disclosed in the document "Xingliang Ma, Qunyu Zhang, Qinlong Zhu, et al. 2015, A Robust CRISPR/Cas9 System for Convenient, High-Efficiency Multiplex Genome Editing in Monocot and Dicot Plants. Molecular Plant, 8(8): 1274-1284".

3. Heat shock method for the introduction of recombinant vector into Agrobacterium tumefaciens EHA105

Referring to the EHA105 competent transformation method, the recombinant plant expression vector pYLCRISPR/Cas9-Pub-H-OsThil-CR constructed above was transformed into the Agrobacterium tumefaciens EHA105 strain by heat shock method. After transformation, a single colony was picked for PCR detection, and the positive clone was named EHA105-OsThil-CRISPR/Cas9-CR.

4. Agrobacterium-mediated rice callus transformation

The specific method of Agrobacterium-mediated rice callus transformation is as follows:

S1. After surface disinfection, rice seeds are inoculated into an induction medium and cultured in a dark place at 30°C for about 1 month. The induced callus tissue is inoculated into a new subculture medium and can be used as material for Agrobacterium transformation after 1 week.

S2. Using the recombinant bacteria prepared above, take out the EHA105-OsThil-CRISPR/Cas9-CR original tube bacterial liquid from the -80℃ refrigerator, streak culture on an LB plate containing 50μg/mL rifampicin and 50μg/mL kanamycin sulfate, and culture in the dark at 28℃ for 2 days. Then, use a pipette to draw 1mL of japonica rice suspension culture medium, wash off the Agrobacterium on the plate, add it to a triangular flask containing 50mL of the suspension, place it in a constant temperature shaker at 28℃ for 1-2h, adjust the OD value to about 1.0, and place it on a clean bench for use;

S3. Select callus tissue with light yellow surface, dry and compact structure, soak it in Agrobacterium suspension for 30 min, then transfer it to solid co-culture medium and co-culture it at 25℃ in the dark for 3 days;

S4. Wash the co-cultured callus tissue with sterile water three times, then dry it with sterile filter paper and transfer it to the screening medium. Culture it in the dark at 30°C. After 20 days, transfer the resistant callus to a new screening medium for a second screening.

S5. The selected resistant callus was transferred to a differentiation medium and cultured in a light incubator at 28°C, 16h light/8h dark, and a light intensity of about 12000LX. After differentiation for about one and a half months, genetically transformed plants (Thil-CR) were obtained.

In the above-mentioned Agrobacterium tumefaciens-mediated genetic transformation of rice mature embryos, the culture medium and formulation components required are shown in Table 4 below.

Table 4 Culture medium

Example 2 Transgenic rice constructed based on recombinant vector overexpressing rice disease susceptibility gene OsThil

1. Design of OsThil target site

The CDS sequence of the OsThil gene (SEQ ID NO. 3) was amplified by PCR and after successful sequencing verification, it was placed in a -20°C refrigerator for future use.

The primer pairs for PCR amplification are:

Thil30-F: 5′-ATGGAGGGAAAGAGGACTACCACTT-3′;

Thil30-R: 5’-TTAGGCTTGTCCTTCCATCTTAGC-3’.

The reaction system is shown in Table 5 below:

Table 5 Amplification reaction system

The reaction program was: 98°C for 5 min; 98°C for 10 s, 55°C for 5 s, 72°C for 5 s, 35 cycles; 72°C for 10 min.

2. Construction of plant expression vector pRHV-OsThil-OE

The CDS sequence of the OsThil gene was constructed into the pRHV vector downstream of the CaMV 35S promoter and upstream of the NOS terminator to obtain the recombinant plant expression vector pRHV-OsThil-OE, the schematic diagram of its T-DNA structure is shown in Figure 4. The above pRHV vector is from the Institute of Crop Sciences, Chinese Academy of Agricultural Sciences.

3. Heat shock method for the introduction of recombinant vector into Agrobacterium tumefaciens EHA105

With reference to the EHA105 competent transformation method, the above-mentioned recombinant plant expression vector pRHV-OsThil-OE was transformed into the Agrobacterium tumefaciens EHA105 strain by heat shock method. After transformation, a single colony was picked and PCR detection was performed using the NosR/HygR primer pair. The positive clone was named pRHV-OsThil-OE (sequence as shown in SEQ ID NO.16).

The reaction system used for PCR detection is shown in Table 6 below:

Table 6 Reaction system

The reaction program was: 98°C for 5 min; 98°C for 10 s, 58°C for 15 s, 72°C for 15 s, 35 cycles; 72°C for 10 min.

4. Agrobacterium-mediated rice callus transformation

S1. After surface disinfection, rice seeds are inoculated into an induction medium and cultured in a dark place at 30°C for about 1 month. The induced callus tissue is inoculated into a new subculture medium and can be used as material for Agrobacterium transformation after 1 week.

S2. Using the recombinant strain prepared above, take out the pRHV-OsThil-OE original tube bacterial solution from the -80°C refrigerator, streak culture on an LB plate containing 50 μg/mL rifampicin and 50 μg/mL kanamycin sulfate, and culture in the dark at 28°C for 2 days. Use a pipette to draw 1 mL of japonica rice suspension culture medium, wash off the Agrobacterium on the plate, add it to a triangular flask containing 50 mL of the suspension, place it on a constant temperature shaker at 28°C for 1-2 hours, adjust the OD value to about 1.0, and place it on a clean bench for use;

S3. Select callus with light yellow surface, dry and compact structure, soak it in Agrobacterium suspension for 30 min, then transfer it to solid co-culture medium and co-culture it at 25℃ in the dark for 3 days;

S4. Wash the co-cultured callus tissue with sterile water three times, then dry it with sterile filter paper and transfer it to the screening medium. Culture it in the dark at 30°C. After 20 days, transfer the resistant callus to a new screening medium for a second screening.

S5. The selected resistant callus was transferred to a differentiation medium and cultured in a light incubator at 28°C, 16h light/8h dark, and a light intensity of about 12000LX. After differentiation for about one and a half months, genetically transformed plants (Thil-OE) were obtained.

The culture medium used above is the same as that in Table 2.

Example 3 CRISPR/Cas9 gene editing of OsThil gene and positive identification of rice mutants overexpressing OsThil gene

1. Positive identification of rice mutants expressing OsThil gene by CRISPR/Cas9 gene editing

The rice leaf DNA was extracted by the CTAB method, and the CRISPR/Cas9 gene-edited OsThil gene rice mutant was amplified by PCR using primer pairs, and the amplified fragment was sequenced for verification (the results are shown in Figure 3, and the deletion sequence is the sequence after the deletion of positions 23-27 (CTTCG, the sequence is shown in SEQ ID NO: 12) or 24-25 (TT) in the sequence shown in SEQ ID NO: 1.).

The primer pairs are as follows:

Thil CR-F: 5′-GGCAGCCATCCACACCTTATGC-3′;

Thil CR-R:5’-CCTCGCTAGATATCAGCTAGGTAGAC-3’.

The amplification reaction system is shown in Table 7 below:

Table 7 Reaction system

The amplification reaction program was: 98°C for 5 min; 98°C for 10 s, 58°C for 15 s, 72°C for 10 s, 35 cycles; 72°C for 10 min.

2. Positive identification of rice mutants overexpressing the OsThil gene

Randomly selected 1/2MS solid medium containing 50 mg·L ^-1 hygromycin for positive identification. The results are shown in Figure 5, showing that the harvested seeds can grow in 1/2MS solid medium containing 50 mg·L ^-1 hygromycin, while the wild type Nipponbare that has not been genetically transformed cannot grow in 1/2MS solid medium containing hygromycin.

Example 3 CRISPR/Cas9 gene editing of OsThil gene and resistance evaluation of rice mutants overexpressing OsThil gene

The roots of rice seedlings cultured in indoor pot experiments can be used for inoculation of root-knot nematodes of the Poaceae family 14 days after sowing. Before inoculation, it is necessary to stop watering with rice complete nutrient solution 1 to 2 days in advance, in order to reduce the moisture in the sand and absorbent resin matrix, so as to facilitate the nematode to infect the rice root system. During inoculation, a blue gun tip is used to insert holes in the sand near the rice stems, and then a rubber-tipped dropper is used to absorb 1 mL of a suspension of about 300 second-instar larvae of the Poaceae root-knot nematodes, and then slowly dripped into the sand and absorbent resin matrix along the holes close to the rice roots. Stop watering with rice complete nutrient solution 2 to 3 days after inoculation to avoid affecting the infection of the root-knot nematodes of the Poaceae family. 14 days after the inoculation of the Poaceae root-knot nematodes, the number of nematode infections and development were counted by acid fuchsin staining, and finally the average value of multiple rice plants was taken for T-test difference analysis (P<0.05).

The results are shown in Figure 7. Compared with Nipponbare rice, overexpression of the OsThil gene in the susceptible variety Nipponbare significantly enhanced the susceptibility to root-knot nematodes of the Poaceae family, while using CRISPR/Cas9 gene editing of the OsThil gene in the susceptible variety Nipponbare significantly enhanced the resistance of rice to root-knot nematodes. Moreover, phenotypic comparison results showed that there was no significant difference between the OsThil mutant plants and the wild-type Nipponbare plants (as shown in Figure 6).

The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims (8)

1. The application of the rice root knot nematode disease gene OsThil in the following (1) or (2):

(1) The application in regulating and controlling the resistance of rice to root-knot nematodes;

(2) Application in cultivating transgenic rice with enhanced susceptibility or resistance to root-knot nematode;

Wherein the nucleotide sequence of the rice root-knot nematode disease gene OsThil is shown as SEQ ID NO. 1.

2. Application of protein expressed by the rice root-knot nematode-susceptibility gene OsThil in culturing transgenic rice with enhanced susceptibility to rice root-knot nematode, wherein the nucleotide sequence of the rice root-knot nematode-susceptibility gene OsThil is shown as SEQ ID NO. 1.

3. Application of a recombinant vector containing a rice root-knot nematode-susceptibility gene OsThil in culturing transgenic rice with enhanced susceptibility to rice root-knot nematodes, wherein the nucleotide sequence of the rice root-knot nematode-susceptibility gene OsThil is shown as SEQ ID NO. 1.

4. The application of a host cell containing a recombinant vector containing a rice root-knot nematode disease gene OsThil in culturing transgenic rice with enhanced susceptibility to rice root-knot nematodes is provided, wherein the nucleotide sequence of the rice root-knot nematode disease gene OsThil is shown as SEQ ID NO. 1.

5. The use according to claim 1, wherein the resistance of rice to root knot nematodes is enhanced by reducing, interfering with the expression of said rice root knot nematode-feel gene OsThil, or knocking out said rice root knot nematode-feel gene OsThil; the rice root-knot nematode disease gene OsThil is overexpressed, so that the susceptibility of the rice to the root-knot nematodes is enhanced.

6. The use according to any one of claims 2 to 4, wherein the susceptibility of rice to root knot nematodes is enhanced by overexpressing the rice root knot nematode disease gene OsThil.

7. A method for cultivating transgenic rice for enhancing susceptibility of rice to root-knot nematodes is characterized by comprising the steps of over-expressing a rice root-knot nematode susceptibility gene OsThil in rice, and cultivating to obtain transgenic rice for enhancing susceptibility of rice to root-knot nematodes; the nucleotide sequence of the rice root-knot nematode disease gene OsThil is shown as SEQ ID NO. 1.

8. A method for cultivating transgenic rice for enhancing the resistance of rice to root-knot nematodes is characterized by comprising the steps of knocking out a rice root-knot nematode-induced gene OsThil from the rice, and cultivating the transgenic rice for enhancing the resistance of the rice to the root-knot nematodes; the nucleotide sequence of the rice root-knot nematode disease gene OsThil is shown as SEQ ID NO. 1.