A gene of brassica napus resistant to sulfonylurea herbicide tribenuron and its application

Abstract

This invention provides a gene for Brassica napus resistant to the sulfonylurea herbicide bensulfuron-methyl, encoding the amino acid sequence shown in SEQ ID NO:2, exhibiting resistance to the sulfonylurea herbicide bensulfuron-methyl. Preferably, the nucleotide sequence of the Brassica napus resistant gene to bensulfuron-methyl is shown in SEQ ID NO:1. The invention also provides the application of the above-mentioned Brassica napus resistant gene to bensulfuron-methyl in detecting plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl, and in cultivating plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl. The Brassica napus resistant gene to bensulfuron-methyl of this invention provides stable resistance to the sulfonylurea herbicide bensulfuron-methyl. Brassica napus with this gene recovers quickly from herbicide damage, exhibits excellent plant type, normal fruit setting, and can be safely used at the recommended field concentration.

Description

Technical Field

[0001] This invention relates to the field of crop genetics and breeding technology, and more specifically, to the field of herbicide-resistant gene technology for Brassica napus, particularly to a gene for Brassica napus resistant to the sulfonylurea herbicide bensulfuron-methyl and its application. Background Technology

[0002] Brassica napus is my country's largest oilseed crop, and rapeseed oil accounts for nearly half of the domestic edible vegetable oil production, making it of great significance to ensuring food and oil security.

[0003] Weeds in the fields consistently reduce rapeseed yields by 10% to 30%, with severe infestations exceeding 50%, making them the primary biological threat restricting yield and quality improvement. Promoting herbicide-resistant varieties and implementing complementary chemical weed control is a core technological path for simplified cultivation, cost reduction, and efficiency improvement. North America and Europe have achieved large-scale application of herbicide-resistant rapeseed, with mainstream types including glyphosate-resistant, glufosinate-resistant, and imidazolinone-resistant varieties. However, the resistance genes for these varieties are restricted by international patent barriers, requiring high patent fees for commercialization; furthermore, glyphosate-resistant and glufosinate-resistant rapeseed are genetically modified varieties. Therefore, creating non-GMO, highly resistant, safe, and herbicide-resistant new germplasm adapted to my country's ecological zones, and cloning independent resistance genes, has significant strategic value for promoting simplified, mechanized, green, and efficient rapeseed production.

[0004] Sulfonylurea (SU) herbicides are the most widely used and developed ultra-efficient herbicides globally, possessing four major advantages: ① Extremely low dosage, requiring only a few grams to tens of grams per hectare; ② Broad spectrum of weed control, effectively controlling broadleaf weeds and partially controlling grassy weeds; ③ Short soil residue period, safe for subsequent crops; ④ Environmentally friendly, with low toxicity to humans and animals. Bensulfuron-methyl, a representative herbicide for wheat fields, is low-cost and has stable efficacy, but its direct application in rapeseed fields can lead to severe phytotoxicity: yellowing of the central leaves, stunting, inhibited bolting, and a sharp drop in seed setting rate. Only bensulfuron-methyl is approved for limited use in rapeseed fields. Currently, rapeseed field weed control mainly relies on pre-emergence herbicides after sowing, resulting in a shortage of broadleaf weed control agents and a narrow safety window during the seedling stage. Breeding rapeseed highly resistant to bensulfuron-methyl allows for its introduction into rapeseed fields, used in combination with monocotyledonous herbicides, significantly broadening the weed control spectrum, reducing weed control costs, and providing a new solution for weed control. However, due to the scarcity of sulfonylurea-resistant rapeseed germplasm with production value, the research and development of commercial sulfonylurea-resistant rapeseed varieties in my country has been very slow, and the creation of independent resistance genes and new germplasm is imminent.

[0005] Acetolactate synthase (ALS, also known as AHAS) is the first key rate-limiting enzyme in the synthesis of branched-chain amino acids (leucine, isoleucine, and valine), and a common target of five major classes of herbicides, including sulfonylureas, imidazolinones (IMI), and triazine (TP). Herbicides kill weeds by binding to ALS, blocking enzyme activity, and inhibiting cell division. The rapeseed genome contains five ALS homologous genes. Among them, BnALS1 and BnALS3 share 98% amino acid homology and are constitutively expressed housekeeping genes, stably expressed in roots, stems, leaves, flowers, and siliques, and are the core functional genes mediating resistance mutations. BnALS2 is specifically expressed only in flowers and siliques; BnALS4 and BnALS5 are pseudogenes. Specific point mutations in the coding region of ALS genes alter the protein conformation, significantly reducing its binding ability to herbicides, thus conferring stable resistance in plants. This is the most mature and widely used molecular mechanism for non-transgenic herbicide-resistant breeding in crops.

[0006] Plant resistance to sulfonamide (SU) herbicides is mainly divided into target resistance (TSR) and non-target resistance (NTSR). Target resistance is caused by ALS point mutations, exhibiting high resistance levels, genetic stability, and significant breeding value, and is the main source of crop resistance. Using Arabidopsis thaliana ALS as a reference, eight key resistance loci have been reported: Ala122, Pro197, Ala205, Asp376, Arg377, Trp574, Ser653, and Gly654. Among them, Pro197, Ala205, and Trp574 contribute the most significantly to SU resistance: the Pro197 mutation usually confers high resistance to SU; the Ala205 mutation provides broad-spectrum resistance to ALS inhibitors; and the Trp574 mutation exhibits extremely high cross-resistance to SU, IMI, and TP. Mutations such as Ser653 mainly mediate resistance to IMI. Non-target resistance depends on metabolic detoxification by P450, GST, etc., which is common in gramineous crops such as rice and wheat, but no natural germplasm suitable for breeding has been found in rapeseed.

[0007] Internationally, Canada primarily uses non-GMO imidazolinone-resistant rapeseed (Clearfield system), with resistance derived from PM1 (BnALS1 Ser653Asp) (Swanson et al. Plant Cell Rep, 1988, 7(2):83-87) and PM2 (BnALS3 Trp574Leu) (Swanson et al. Theor Appl Genet, 1989, 78(4):525-530; Swanson et al. 1989; Hattori et al. Mol Gen Genet, 1995, 246:419-425), obtained through microspore mutagenesis, and already industrialized. Australia also mainly uses non-GMO imidazolinone-resistant rapeseed. All of the above materials target imidazolinone resistance, exhibiting limited or no resistance to bensulfuron-methyl, and their ecological adaptability is not compatible with my country's main rapeseed-producing areas.

[0008] Domestic research on herbicide-resistant rapeseed has made phased progress: The team led by Pu Huiming at the Jiangsu Academy of Agricultural Sciences screened the natural mutant M9 (Gao Jianqin et al., Journal of Plant Genetic Resources, 2010, 11(3):369-373), and obtained M343 (ZL201310111739.5) through EMS mutagenesis, with resistances of BnALS1 Ser653Asp and BnALS3 Trp574Leu, respectively; 12WH318 from the Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences (Li Xiaotong et al., Chinese Journal of Oil Crops, 2015, 37(3):269-276) also has BnALS1 Ser653Asp; Huazhong Agricultural University obtained the BnALS3 Pro197Ser mutant from “Huashuang 5” (Li et al. Theor Appl.). Genet, 2015, 128(1):107-118); The team led by Hu Shengwu of Northwest A&F University obtained the K5 mutant (CN106754997A) from "Zhongshuang 9", which is BnALS1 Pro182Ser (C544T) and has certain resistance to bensulfuron-methyl. The above research laid the foundation for herbicide-resistant rapeseed breeding in my country, but there are still obvious shortcomings: ① Most materials are mainly resistant to IMI, and there are few independent germplasm resistant to bensulfuron-methyl; ② The resistance of introduced or reported mutants is unstable and the agronomic traits deteriorate in the high humidity, low sunshine and rainy ecological zone of Southwest China; ③ The resistance effect, safety and adaptability of different sites of the same gene and different base mutations at the same site are significantly different. Existing genes are difficult to meet the needs of the Southwest production area for "high resistance, safety, fast recovery and good traits". It is still necessary to explore new, specific and independent intellectual property rights ALS resistance sites.

[0009] Therefore, it is desirable to provide a gene for Brassica napus resistant to the sulfonylurea herbicide bensulfuron-methyl, which is resistant to bensulfuron-methyl with stable resistance. Brassica napus with this gene recovers quickly from herbicide damage, has excellent plant type, and normal fruit setting, and can be safely used at the recommended concentration in the field. Summary of the Invention

[0010] In order to overcome the shortcomings of the prior art, one object of the present invention is to provide a gene for rapeseed that is resistant to the sulfonylurea herbicide bensulfuron-methyl. This gene is resistant to the sulfonylurea herbicide bensulfuron-methyl, has stable resistance, and rapeseed with this gene recovers quickly from herbicide damage, has excellent plant type, and normal fruit setting. It can be safely used in the field at the recommended concentration and is suitable for large-scale promotion and application.

[0011] Another objective of this invention is to provide an application of a gene for rapeseed resistant to the sulfonylurea herbicide bensulfuron-methyl, which can be used to detect plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl. This method is simple and quick, so as to determine whether the plant can be treated with the sulfonylurea herbicide bensulfuron-methyl, and is suitable for large-scale promotion and application.

[0012] Another objective of this invention is to provide an application of a gene for rapeseed resistant to the sulfonylurea herbicide bensulfuron-methyl, which can be used to cultivate plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl. This resistant plant exhibits stable resistance, rapid recovery from herbicide damage, excellent plant type, and normal fruit setting. It can be safely applied at the recommended concentration in the field and is suitable for large-scale promotion and application.

[0013] To achieve the above objectives, in a first aspect of the present invention, a gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in Brassica napus is provided, characterized in that the amino acid sequence encoded by the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in Brassica napus is shown in SEQ ID NO: 2, and the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in Brassica napus possesses resistance to the sulfonylurea herbicide bensulfuron-methyl.

[0014] Preferably, the nucleotide sequence of the gene for the rapeseed resistant to the sulfonylurea herbicide bensulfuron is shown in SEQ ID NO: 1.

[0015] In a second aspect of the invention, the application of the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in the above-mentioned rapeseed is provided in the detection of plants, plant cells or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl.

[0016] Preferably, detection primers are designed based on the gene of Brassica napus resistant to the sulfonylurea herbicide bensulfuron-methyl. The detection primers are used to perform PCR amplification on plants, plant cells or plant tissues to obtain PCR products. The PCR products are then sequenced to determine whether the plant, plant cells or plant tissues are plants, plant cells or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl.

[0017] Preferably, the plant is Brassica napus, the plant cells are Brassica napus cells, and the plant tissue is Brassica napus tissue.

[0018] In a third aspect of the invention, the application of the above-mentioned gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in the cultivation of plants, plant cells or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl is provided.

[0019] Preferably, the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in Brassica napus is introduced into the plant, plant cell, or plant tissue to obtain the plant, plant cell, or plant tissue resistant to the sulfonylurea herbicide bensulfuron-methyl.

[0020] Preferably, the plant is Brassica napus, the plant cells are Brassica napus cells, and the plant tissue is Brassica napus tissue.

[0021] The beneficial effects of this invention are as follows:

[0022] 1. The amino acid sequence encoded by the gene for resistance to the sulfonylurea herbicide bensulfuron in Brassica napus of the present invention is shown in SEQ ID NO: 2. The gene for resistance to the sulfonylurea herbicide bensulfuron in Brassica napus contains resistance to the sulfonylurea herbicide bensulfuron. Therefore, it can resist the sulfonylurea herbicide bensulfuron, and the resistance is stable. Brassica napus with this gene recovers quickly from herbicide damage, has excellent plant type, and normal fruit setting. It can be safely used at the recommended concentration in the field and is suitable for large-scale promotion and application.

[0023] 2. The application of the gene for sulfonylurea-resistant bensulfuron-methyl in the detection of plants, plant cells, or plant tissues resistant to bensulfuron-methyl in the present invention is simple and quick. Therefore, it can be used to detect plants, plant cells, or plant tissues resistant to bensulfuron-methyl, so as to determine whether bensulfuron-methyl can be used for weed control. It is suitable for large-scale promotion and application.

[0024] 3. The application of the above-mentioned gene for resistance to sulfonylurea herbicide bensulfuron in the cultivation of plants, plant cells or plant tissues resistant to sulfonylurea herbicide bensulfuron in the present invention. Therefore, it can be used to cultivate plants, plant cells or plant tissues resistant to sulfonylurea herbicide bensulfuron, which can resist sulfonylurea herbicide bensulfuron, have stable resistance, recover quickly from herbicide damage, have excellent plant type, and normal fruit setting. It can be safely used at the recommended concentration in the field and is suitable for large-scale promotion and application.

[0025] These and other objects, features and advantages of the present invention will be fully apparent from the following detailed description and drawings, and can be achieved by the methods, means and combinations thereof specifically pointed out in the specification. Attached Figure Description

[0026] Figure 1 These are phenotypic images of GR724R resistance identification in Brassica napus, where 1 is the negative control and 2 is GR724R.

[0027] Figure 2 Gel electrophoresis images of the amplification products obtained by PCR amplification of DNA from Brassica napus GR724R. In the image, 1 is the DNA ladder (Vazyme, DL2000 Plus DNA Marker), in bp; 2 is the amplification product.

[0028] Figure 3 This is a partial image showing the alignment results of the full-length BnALS1G1676T gene sequence with the full-length BnALS1 gene sequence. The T bases enclosed in the box are mutated bases. Detailed Implementation

[0029] To address the prominent issue of my country's reliance on imported herbicide-resistant rapeseed germplasm resources and the lack of genes with independent intellectual property rights, the inventors conducted radiation mutagenesis and rigorous multi-generation screening of bensulfuron-methyl in Brassica napus, obtaining a new non-transgenic germplasm GR724R resistant to the sulfonylurea herbicide bensulfuron-methyl. This led to the discovery of a gene in Brassica napus resistant to bensulfuron-methyl, which exhibits stable resistance. Brassica napus with this gene recovers quickly from herbicide damage, displays excellent plant architecture, and produces normal seeds, providing a new genetic resource for herbicide-resistant rapeseed breeding. Based on this, the present invention was completed.

[0030] This invention first provides a gene for rapeseed resistant to the sulfonylurea herbicide bensulfuron-methyl. The amino acid sequence encoded by the gene is shown in SEQ ID NO: 2. The gene for rapeseed resistant to the sulfonylurea herbicide bensulfuron-methyl exhibits resistance to the sulfonylurea herbicide bensulfuron-methyl.

[0031] The gene for the rapeseed resistant to the sulfonylurea herbicide bensulfuron can have any suitable nucleotide sequence. Preferably, the nucleotide sequence of the rapeseed resistant to the sulfonylurea herbicide bensulfuron is shown in SEQ ID NO: 1.

[0032] This invention also provides the application of the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in Brassica napus in detecting plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron-methyl.

[0033] The detection of plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron can be performed using any suitable method. Preferably, detection primers are designed based on the gene of the Brassica napus resistant to the sulfonylurea herbicide bensulfuron. The detection primers are used to perform PCR amplification on the plants, plant cells, or plant tissues to obtain PCR products. The PCR products are then sequenced to determine whether the plants, plant cells, or plant tissues are resistant to the sulfonylurea herbicide bensulfuron.

[0034] The plant, plant cell, or plant tissue can be any suitable plant, plant cell, or plant tissue. Preferably, the plant is Brassica napus, the plant cell is Brassica napus cell, and the plant tissue is Brassica napus tissue.

[0035] The present invention also provides the application of the above-mentioned gene for resistance to the sulfonylurea herbicide bensulfuron in rapeseed in the cultivation of plants, plant cells or plant tissues resistant to the sulfonylurea herbicide bensulfuron.

[0036] The cultivation of plants, plant cells, or plant tissues resistant to the sulfonylurea herbicide bensulfuron can be achieved by any suitable method. Preferably, the gene for bensulfuron resistance in Brassica napus is introduced into the plant, plant cell, or plant tissue to obtain the plant, plant cell, or plant tissue resistant to the sulfonylurea herbicide bensulfuron.

[0037] The plant, plant cell, or plant tissue can be any suitable plant, plant cell, or plant tissue. Preferably, the plant is Brassica napus, the plant cell is Brassica napus cell, and the plant tissue is Brassica napus tissue.

[0038] To provide a clearer understanding of the technical content of this invention, the following embodiments are provided for detailed description. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions or as recommended by the manufacturer. Reagents or instruments used, unless otherwise specified, are all commercially available products.

[0039] Example 1: Screening of GR724R, a mutant of Brassica napus resistant to the sulfonylurea herbicide bensulfuron-methyl.

[0040] 1. Experimental material: Brassica napus GR724 (provided by Guizhou Crop Germplasm Bank, available to the general public upon application).

[0041] 2. Mutagenic treatment: 2000 Gy dose 60 20 kg of dry seeds of Brassica napus GR724 were treated with Co-γ ray irradiation and directly planted to obtain M1 generation plants. When the seedlings were 4-6 leaves, 1.5 g / mu of bensulfuron-methyl active ingredient was sprayed uniformly. For the plants that were not killed, they were self-pollinated and bagged to obtain M2 generation seeds.

[0042] 3. Field sowing: M2 generation seeds were sown at the Guiyang Experimental Base of Guizhou Academy of Agricultural Sciences. When the seedlings were 4-6 leaves, 1.5g / mu of bensulfuron-methyl active ingredient was sprayed uniformly.

[0043] 4. Phenotypic identification and screening:

[0044] Phenotypic identification is shown in Figure 1 As shown.

[0045] The resistant material with the best resistance to bensulfuron-methyl: after spraying bensulfuron-methyl, only the central leaves turn slightly yellow, and they can naturally recover and turn green in 7-10 days. The plants continue to grow normally, without stunting or dying, and the growth process is normal. They can successfully bolt, bud, flower, and bear fruit. They have excellent agronomic traits and are named GR724R.

[0046] Seeds of GR724 that have not undergone radiation mutagenesis were directly sown to obtain seedlings of the same period as the control: all plants were severely yellowed, wilted, and died, and could not grow normally.

[0047] 5. Strain purification: GR724R seeds are harvested from individual plants and self-pollinated for two generations to obtain a stable genetically resistant bensulfuron-methyl GR724R strain, which is then preserved in the Guizhou Provincial Crop Germplasm Bank and is available to the public upon application.

[0048] Example 2 Cloning and Sequence Analysis of the BnALS1G1676T Gene

[0049] DNA extraction: The seeds of the bensulfuron-resistant GR724R strain from Example 1 and the seeds of wild-type GR724 were sown at the Guiyang Experimental Base of the Guizhou Academy of Agricultural Sciences. Young leaves were taken and high-quality DNA was extracted using the CTAB method.

[0050] Specific PCR amplification: Based on the conserved region of the ALS1 gene in Brassica napus (reference sequence Z11524.1, https: / / www.ncbi.nlm.nih.gov / nuccore / Z11524.1), specific primers were designed (forward primer as shown in SEQ ID NO: 3, reverse primer as shown in SEQ ID NO: 4). PCR amplification was performed (PCR amplification system: 50 ng DNA template, 1 μl each of approximately 10 mM primers, 25 μl of 2×Phanta Max Master Mix, and ddH2O added to 50 μl; PCR amplification program: 95℃ for 3 min, (95℃ for 15 s, 56℃ for 15 s, 72℃ for 1 min) × 33 cycles, 72℃ for 5 min). The PCR products were obtained; please refer to [link to relevant documentation]. Figure 2 As shown.

[0051] Cloning and sequencing: The PCR product was ligated into the pMD18-T cloning vector, transformed into E. coli, and positive clones were sent for sequencing to obtain the full-length gene sequence of BnALS1G1676T of GR724R and the full-length gene sequence of BnALS1 of GR724.

[0052] Sequence alignment: The full-length BnALS1G1676T gene sequence was aligned with the full-length BnALS1 gene sequence of GR724, i.e., the ALS1 gene sequence of wild-type GR724. For partial alignment results, please refer to [link to relevant documentation]. Figure 3 As shown, it was determined that BnALS1G1676T has a specific resistance mutation site, which is the 1676th base of the ALS1 gene, where G is mutated to T, resulting in the 559th amino acid of its coding sequence being mutated from tryptophan (W) to leucine (L).

[0053] Example 3: Cultivation of herbicide-resistant rapeseed

[0054] Recipient material: GRD328, the paternal parent of Youyan 200, a high-yield, high-quality, and disease-resistant rapeseed variety promoted in Guizhou Province (provided by the Guizhou Crop Germplasm Bank, available to any member of the public upon application).

[0055] Breeding method: GR724R is used as the donor and crossed with the recipient parent GRD328, followed by continuous backcrossing.

[0056] Resistance screening: Spray 1.5g / mu of bensulfuron-methyl at the 4-6 leaf stage of the offspring seedlings to obtain surviving plants, which are resistant individual plants.

[0057] Molecular identification: Following the method in Example 2, PCR amplification / sequencing was performed using the specific primers from Example 2 to confirm the stable integration of the BnALS11676T gene in the resistant single plant.

[0058] Example 4 Field Application

[0059] Hybrid seeds were prepared by using GR724R and the female parent GR722 of the rapeseed hybrid variety Youyan 300 (provided by the Guizhou Provincial Crop Germplasm Bank and available to the public upon application). Applying a low concentration of bensulfuron-methyl (5-8 PPM) can prevent the female parent of Youyan 300 from being killed but still produce male sterility. The growth of the male parent GR724R is not affected by bensulfuron-methyl. The sulfonylurea herbicide bensulfuron-methyl can be sprayed on both the male and female parents at the seed production site simultaneously, eliminating the need for protection of the male parent. This reduces labor and improves seed production efficiency.

[0060] The field application specifications for Brassica napus containing the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl of this invention are as follows:

[0061] Optimal application period: rapeseed at 4-6 leaf stage, weeds at 2-4 leaf stage;

[0062] Recommended dosage: bensulfuron-methyl 1.5g / mu;

[0063] Efficacy: Total weed control efficacy ≥90%, no herbicide damage to rapeseed, and normal yield increase of 8%~15%.

[0064] The gene BnALS1G1676T for resistance to the sulfonylurea herbicide bensulfuron-methyl in Brassica napus of this invention is a mutant gene obtained by radiation-induced specific point mutation in the acetolactate synthase (ALS1) encoding gene. Alternatively, it can be obtained through conventional molecular biology methods in the art, such as gene synthesis, PCR amplification, and site-directed mutagenesis. This gene endows Brassica napus with stable, high-level, and safe resistance to bensulfuron-methyl, possessing the following core characteristics:

[0065] (1) Constitutive stable expression: It is expressed in roots, stems, leaves, flowers and siliques, with the highest expression level in leaves;

[0066] (2) Safety of resistance concentration: It is safe for rapeseed at a concentration of 1.0~2.0 g / mu of bensulfuron-methyl, and the recommended concentration for field is 1.5 g / mu;

[0067] (3) No adverse chain reaction burden: GR724R has excellent plant type, lodging resistance, disease resistance, seed setting rate, oil content and other traits;

[0068] (4) Stable inheritance: The resistance trait follows Mendelian inheritance laws and can be stably passed on to offspring through hybridization and backcrossing.

[0069] By using GR724R as the resistance donor parent and crossing it with superior rapeseed varieties through hybridization and backcrossing, new rapeseed varieties resistant to bensulfuron can be bred.

[0070] Compared with the prior art, the present invention has the following advantages:

[0071] (1) Complete independent intellectual property rights: The gene nucleotide sequence and amino acid sequence are both disclosed for the first time and have not been seen in any patents or literature reports. This fills the gap in bensulfuron-resistant rapeseed gene resources in Southwest my country and has important breeding value and industrial prospects.

[0072] (2) Strong resistance and high safety: At the recommended concentration, only the heart leaves turn slightly yellow and can recover quickly, and there is no lethal phytotoxicity to rapeseed;

[0073] (3) Highly adapted to the ecology of Southwest China: It is tolerant to moisture and low light, and has good early maturity, making it suitable for planting in Guizhou and Southwest China;

[0074] (4) Excellent agronomic traits: It does not affect yield, quality, disease resistance, or lodging resistance, and can be directly used as a parent for breeding;

[0075] (5) Significantly reduce costs and increase efficiency: One application during the seedling stage can effectively control weeds, reducing labor input by more than 60%;

[0076] (6) High breeding efficiency: It can be quickly tracked through molecular markers to achieve rapid aggregation of resistance and excellent agronomic traits.

[0077] This invention uses GR724, a type of rapeseed, as the original material. An M2 generation mutant population was obtained through radiation mutagenesis. Resistance was assessed by spraying with an effective concentration of 1.5 g / mu of bensulfuron-methyl at the 4-6 leaf stage. The optimal herbicide-resistant material, GR724R, was selected. A bensulfuron-resistant gene was cloned from GR724R. This gene encodes acetolactate synthase (ALS1), and compared to wild-type GR724, its coding region contains a specific resistance mutation site, stably conferring high-level resistance to bensulfuron-methyl in rapeseed. After spraying with the recommended concentration of bensulfuron-methyl, GR724R only showed slight yellowing of the central leaves, which later naturally turned green. It could complete normal growth, bolting, flowering, and fruiting without any risk of phytotoxicity. This gene consists of 1965 bases, with the nucleotide sequence SEQ ID NO: 1. The protein sequence obtained by translating this gene is SEQ ID NO: 2. Introducing this gene into plants resistant to bensulfuron-methyl conjugates resistance to the herbicide bensulfuron-methyl, thereby enhancing the plant's resistance to bensulfuron-methyl. This invention provides a new gene and germplasm with completely independent intellectual property rights for simplified and mechanized cultivation of rapeseed in my country, which can significantly reduce the labor costs of weeding in the field and improve rapeseed production efficiency and planting benefits.

[0078] In summary, the gene for resistance to the sulfonylurea herbicide bensulfuron-methyl in the present invention can resist the sulfonylurea herbicide bensulfuron-methyl with stable resistance. Brassica napus with this gene recovers quickly from herbicide damage, has excellent plant type, and normal fruit setting. It can be safely used in the field at the recommended concentration and is suitable for large-scale promotion and application.

[0079] In this specification, the invention has been described with reference to specific embodiments thereof. However, it will be apparent that various modifications and variations can be made without departing from the spirit and scope of the invention. Therefore, the specification and drawings should be considered illustrative rather than restrictive.

Claims

1. A Brassica napus gene for resistance to the sulfonylurea herbicide tribenuron, characterized in that, The amino acid sequence of the Brassica napus gene resistant to the sulfonylurea herbicide tribenuron is shown as SEQ ID NO: 2, and the Brassica napus gene resistant to the sulfonylurea herbicide tribenuron has the resistance to the sulfonylurea herbicide tribenuron.

2. The Brassica napus gene for resistance to the sulfonylurea herbicide tribenuron according to claim 1, characterized in that, The nucleotide sequence of the Brassica napus gene resistant to the sulfonylurea herbicide tribenuron is shown as SEQ ID NO:

1.

3. The Brassica napus gene resistant to the sulfonylurea herbicide tribenuron of claim 1 is applied to detect plants, plant cells or plant tissues resistant to the sulfonylurea herbicide tribenuron.

4. Use according to claim 3, characterized in that, According to the Brassica napus gene resistant to the sulfonylurea herbicide tribenuron, a detection primer is designed, and the detection primer is used for PCR amplification of plants, plant cells or plant tissues to obtain a PCR product, and the PCR product is sequenced to determine whether the plants, plant cells or plant tissues are the plants, plant cells or plant tissues resistant to the sulfonylurea herbicide tribenuron.

5. Use according to claim 3, characterized in that, The plants are Brassica napus, the plant cells are Brassica napus cells, and the plant tissues are Brassica napus tissues.

6. The Brassica napus gene resistant to the sulfonylurea herbicide tribenuron of claim 1 is applied to breed plants, plant cells or plant tissues resistant to the sulfonylurea herbicide tribenuron.

7. Use according to claim 6, characterized in that, The Brassica napus gene resistant to the sulfonylurea herbicide tribenuron is introduced into the plants, plant cells or plant tissues to obtain the plants, plant cells or plant tissues resistant to the sulfonylurea herbicide tribenuron.

8. Use according to claim 6, characterized in that, The plants are Brassica napus, the plant cells are Brassica napus cells, and the plant tissues are Brassica napus tissues.

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