Low glutelin high dietary fiber excellent flavor rice and breeding method and application thereof

Abstract

The application provides a low-gluten high-dietary fiber excellent flavor rice as well as a breeding method and application thereof, and relates to the technical field of rice breeding. The breeding method uses a high-dietary fiber rice variety as a female parent and a low-gluten flavor rice variety as a male parent to perform hybridization, and then performs genotype detection on the F2 generation by using Badh-M2 and Lgc1 molecular markers respectively, and combines with agronomic trait selection to realize efficient polymerization of three functional genes of flavor, low gluten and high dietary fiber. The method significantly improves the selection efficiency, avoids trait loss and cycle extension caused by multiple backcrossing. The bred rice has the effects of rich flavor, significantly reduced rice gluten content, obviously increased dietary fiber content and reduced blood glucose generation index, and meanwhile, maintains the effects of high yield, early maturity, excellent plant leaf morphology and good ecological adaptability, and can meet the nutritional needs of chronic kidney disease, diabetes and obese people for functional staple food.

Description

Technical Field

[0001] This invention relates to the field of rice cultivation technology, and in particular to a low-gluten, high-dietary-fiber, high-fragrant rice variety, its cultivation method, and its application. Background Technology

[0002] With the rapid development of society and economy and the improvement of people's living standards, people's eating habits have changed. Due to long-term overeating and a habit of consuming high-salt, high-fat, and high-calorie foods, the incidence of chronic diseases such as hyperglycemia, hyperlipidemia, hypertension, diabetes, cardiovascular disease, and chronic kidney disease has continued to rise.

[0003] For example, excessive intake of absorbable protein can burden the kidneys and worsen the condition in patients with chronic kidney disease. Therefore, a low-protein diet is an important treatment for chronic kidney disease patients. Gluten is the most abundant protein stored in rice seeds and is easily absorbed by the body. Low-gluten rice, with a gluten content of less than 4%, can be used as a staple food for patients with chronic kidney disease during their rehabilitation. Furthermore, dietary fiber is not digested by the gastrointestinal tract and does not produce energy. It has advantages such as controlling glycemic index, preventing type II diabetes, increasing satiety, controlling weight, preventing lipid metabolism disorders, and preventing colon cancer. Therefore, high-fiber rice is an excellent choice for diabetic patients and obese individuals whose staple food is rice.

[0004] Therefore, cultivating rice varieties that are low in gluten and high in dietary fiber is an effective way to meet the dietary needs of many patients with chronic kidney disease and diabetes. However, existing rice varieties generally have the problem of high gluten content (about 8-12%) and low dietary fiber content (about 0.4-0.7%), making it difficult to balance multiple nutritional functions and eating quality.

[0005] Furthermore, in breeding practice, the three traits of "fragrance", "low gluten content" and "high dietary fiber" are controlled by different genes, and there is a lack of efficient prospect screening methods. Currently, there is a lack of excellent rice varieties on the market that simultaneously possess these three genes.

[0006] In view of this, the present invention is hereby proposed. Summary of the Invention

[0007] The primary objective of this invention is to provide a method for cultivating a high-quality aromatic rice with low gluten content and high dietary fiber. This method achieves efficient integration of three major functional characteristics: low gluten content, high dietary fiber, and aroma, significantly improving breeding selection efficiency and ultimately obtaining an aromatic rice variety, "Jiugu No. 2," which possesses low gluten content, high dietary fiber, and a low glycemic index. Furthermore, this variety exhibits high yield, early maturity, excellent plant and leaf morphology, and broad ecological adaptability.

[0008] The second objective of this invention is to provide a low-gluten, high-dietary-fiber rice with excellent aroma.

[0009] The third objective of this invention is to provide an application of a low-gluten, high-dietary-fiber rice with excellent aroma.

[0010] In order to achieve the above-mentioned objectives of the present invention, the following technical solution is adopted: This invention provides a method for cultivating a low-gluten, high-dietary-fiber, high-aroma rice variety, the cultivation method comprising: By using hybridization, rice varieties with high dietary fiber were used as the female parent to cross with fragrant rice varieties. The hybrid offspring were then subjected to marker-assisted selection using DNA molecular markers, combined with agronomic trait selection, to obtain high-quality fragrant rice with low gluten content and high dietary fiber. The marker-assisted selection begins in the F2 generation, and the markers include Badh-M2 and Lgc1 markers.

[0011] Furthermore, the molecular marker-assisted selection includes: During the F2 generation rice seedling stage, the genotype of individual plants was detected using Badh-M2 and Lgc1 molecular markers to screen out individual plants that were homozygous for both the Badh2 aroma gene and the Lgc1 low glutenin gene. Then, the seeds of the planted individual plants were soaked in water and placed under a transmission optical detection device. Based on the light transmittance of the seeds, plump, opaque, chalky seeds were selected.

[0012] Furthermore, the seeds are soaked in water for 3-5 hours.

[0013] Furthermore, the primer sequences for the Badh-M2 molecular marker are shown in SEQ ID NO. 1 and SEQ ID NO. 2; The sequence of SEQ ID NO. 1 is as follows: 5'-TAGGTTGCATTTACTGGGAG-3'; The sequence of SEQ ID NO. 2 is as follows: 5'-GGAAACAAACCTTAACCATAG-3'.

[0014] Furthermore, the Lgc1 molecular marker includes the Lgc1-1 primer sequence and the Lgc1-2 primer sequence; The Lgc1-1 primer sequences are shown in SEQ ID NO. 3 and SEQ ID NO. 4; The sequence of SEQ ID NO. 3 is as follows: 5'-TTCTACAATGAAGGCGATGC-3'; The sequence of SEQ ID NO. 4 is as follows: 5'-CTGGGCTTTAACGGGACT-3'; The Lgc1-1 primer sequences are shown in SEQ ID NO. 5 and SEQ ID NO. 6; The sequence of SEQ ID NO. 5 is as follows: 5'-ACCGTGTTATGGCAGTTT-3'; The sequence of SEQ ID NO. 6 is as follows: 5'-ATTCAAGGGCTATCGTCT-3'.

[0015] Furthermore, the reaction system for the molecular marker-assisted selection PCR amplification is 10 μL, comprising: 1 μL template DNA, 1 μL 10×PCR Buffer, 1 μL 2 mmol / L dNTP, 1 μL primer combination, 0.2 μL Taq DNA polymerase, and the remainder being sterile deionized water. The PCR amplification program for the Badh-M2 label is as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 20 s, 35 cycles; 72℃ final extension for 5 min. The PCR amplification program for the Lgc1 label is as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 50 s, 35 cycles; 72℃ final extension for 5 min.

[0016] Furthermore, the cultivation method includes: S1: A hybrid F1 is obtained by crossing a rice variety with high dietary fiber as the female parent with a rice variety with aroma. S2: Plant F1, remove false hybrids, and harvest the seeds after maturity; S3: Plant F2, and use aroma gene linkage markers and low gluten protein gene linkage markers to perform genotyping on individual plants during the seedling stage, and screen out individual plants that are homozygous for both aroma gene and low gluten protein gene, and harvest individual plants for seed. S4: After soaking the seeds obtained in S3 in water, place them under a transmission optical detection device and screen for plump, opaque, chalky white seeds based on their light transmittance to determine them as high-dietary-fiber individuals. S5: Plant the seeds obtained from S4 screening, select the best single plants based on agronomic traits, and harvest seeds from a single plant with a single ear. The entire growth period of the superior single plant is 110-120 days, and its entire growth period is ≤ that of the rice variety Tianyou Huazhan; the number of effective tillers of the single plant is ≥8; S6: Plant the seeds obtained in S5, and repeat steps S4 and S5 to select seeds with good agronomic traits; S7: Seeds obtained from planting S6 are continuously self-pollinated until the phenotype is stable, and seeds are harvested from a single plant. S8: Seeds obtained from planting S7 were planted in plots and variety comparison trials were conducted. At the same time, the gluten content and dietary fiber content in the rice were tested. After comparison, strains with stable phenotypes, early growth period, high yield, low gluten content and high dietary fiber content were selected to complete the breeding.

[0017] It should be noted that "high dietary fiber" in this application refers to rice varieties or individuals with high dietary fiber content, and "high dietary fiber content" means that the refined rice has a dietary fiber content of ≥4.0%.

[0018] This invention provides a low-gluten, high-dietary-fiber, and aromatic rice variety cultivated according to the above-described cultivation method.

[0019] Furthermore, the rice has a gluten content of <4%, a dietary fiber content of ≥4.0%, and a rice aroma; The glycemic index (GI) of the rice is ≤55.

[0020] The use of the above-mentioned low-gluten, high-dietary-fiber, and aromatic rice provided by this invention in the preparation of functional staple food products; The functional staple food mentioned above is a functional staple food for patients with chronic kidney disease, diabetes, or obesity.

[0021] Compared with the prior art, the beneficial effects of the present invention are as follows: This invention relates to a method for cultivating a high-quality aromatic rice with low gluten content and high dietary fiber. The method involves crossing a high-dietary-fiber maternal parent with a low-gluten content aromatic parent, selecting agronomic traits in the F2 generation, and combining gene aggregation with molecular marker-assisted selection technology. Pedigree selection and pressure screening are used to improve selection efficiency. The resulting rice variety has a fragrant aroma, significantly reduced gluten content, significantly increased dietary fiber content, and a lower glycemic index. It also exhibits high yield, early maturity, excellent plant and leaf morphology, and wide ecological adaptability.

[0022] The low-gluten, high-fiber, and high-tasting rice provided by this invention can be widely used in the preparation of functional staple food products, which are intended for patients with chronic kidney disease, diabetes, or obesity. Attached Figure Description

[0023] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0024] Figure 1 This is a technical roadmap for the cultivation of the low-gluten, high-dietary-fiber aromatic rice variety "Jiugu No. 2" provided in Example 1 of the present invention. Figure 2 This is a graph showing the electrophoresis results of the F2 generation Badh-M2 molecular marker PCR amplification provided in Example 1 of this invention; Figure 3 This is a graph showing the electrophoresis results of Lgc1 molecular marker PCR amplification in the F2 generation population provided in Example 1 of this invention. Figure 4 This is a field phenotypic diagram of a typical single plant of Jiugu No. 2 provided in Example 1 of the present invention; Figure 5 This is an appearance diagram of Jiugu No. 2 polished rice provided in Experimental Example 1 of the present invention; Figure 6 The image shows the SDS-PAGE electrophoresis pattern of total protein from Jiugu No. 2 rice provided in Experimental Example 1 of this invention. Detailed Implementation

[0025] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0026] According to one aspect of the present invention, a method for cultivating a low-gluten, high-dietary-fiber, aromatic rice variety, the method comprising: By using hybridization, rice varieties with high dietary fiber were used as the female parent to cross with fragrant rice varieties. The hybrid offspring were then subjected to marker-assisted selection using DNA molecular markers, combined with agronomic trait selection, to obtain high-quality fragrant rice with low gluten content and high dietary fiber. The marker-assisted selection begins in the F2 generation, and the markers include Badh-M2 and Lgc1 markers.

[0027] This invention relates to a method for cultivating a high-quality aromatic rice with low gluten content and high dietary fiber. The method involves crossing a high-dietary-fiber maternal parent with a low-gluten content aromatic parent, selecting agronomic traits in the F2 generation, and combining gene aggregation with molecular marker-assisted selection technology. Pedigree selection and pressure screening are used to improve selection efficiency. The resulting rice variety has a fragrant aroma, significantly reduced gluten content, significantly increased dietary fiber content, and a lower glycemic index. It also exhibits high yield, early maturity, excellent plant and leaf morphology, and wide ecological adaptability.

[0028] In a preferred embodiment of the present invention, the molecular marker-assisted selection includes: During the F2 generation rice seedling stage, the genotype of individual plants was detected using Badh-M2 and Lgc1 molecular markers to screen out individual plants that were homozygous for both the Badh2 aroma gene and the Lgc1 low glutenin gene. Then, the seeds of the planted individual plants were soaked in water and placed under a transmission optical detection device. Based on the light transmittance of the seeds, plump, opaque, chalky seeds were selected.

[0029] As a preferred embodiment, this invention simultaneously detects Badh-M2 and Lgc1 molecular markers during the F2 generation seedling stage, which can accurately screen out single plants that are homozygous for both aroma and low glutenin genes, thereby achieving the aggregation of bifunctional genes at an early stage. Combined with transmission optical detection after water immersion, it can effectively identify high chalky seeds, providing a rapid and non-destructive screening basis for high dietary fiber traits, and significantly improving the accuracy and efficiency of multi-trait aggregation.

[0030] In a preferred embodiment of the present invention, the seeds are soaked in water for 3 to 5 hours, preferably 3 hours.

[0031] As a preferred embodiment, the present invention optimizes the endosperm water absorption state by controlling the seed water soaking time to 3-5 hours, making the chalky white features of the seeds clearer and more stable in subsequent transmission optical detection, thereby improving the accuracy and repeatability of high dietary fiber candidate seed identification.

[0032] In a preferred embodiment of the present invention, the primer sequences of the Badh-M2 molecular marker are shown in SEQ ID NO. 1 and SEQ ID NO. 2; The sequence of SEQ ID NO. 1 is as follows: 5'-TAGGTTGCATTTACTGGGAG-3'; The sequence of SEQ ID NO. 2 is as follows: 5'-GGAAACAAACCTTAACCATAG-3'.

[0033] In a preferred embodiment of the present invention, the Lgc1 molecular marker includes an Lgc1-1 primer sequence and an Lgc1-2 primer sequence; The Lgc1-1 primer sequences are shown in SEQ ID NO. 3 and SEQ ID NO. 4; The sequence of SEQ ID NO. 3 is as follows: 5'-TTCTACAATGAAGGCGATGC-3'; The sequence of SEQ ID NO. 4 is as follows: 5'-CTGGGCTTTAACGGGACT-3'; The Lgc1-1 primer sequences are shown in SEQ ID NO. 5 and SEQ ID NO. 6; The sequence of SEQ ID NO. 5 is as follows: 5'-ACCGTGTTATGGCAGTTT-3'; The sequence of SEQ ID NO. 6 is as follows: 5'-ATTCAAGGGCTATCGTCT-3'.

[0034] In a preferred embodiment of the present invention, the reaction system of the molecular marker-assisted selection PCR amplification is 10 μL, comprising: 1 μL template DNA, 1 μL 10×PCR Buffer, 1 μL 2 mmol / L dNTP, 1 μL primer combination, 0.2 μL Taq DNA polymerase, and the remainder is sterile deionized water. The PCR amplification program for the Badh-M2 label is as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 20 s, 35 cycles; 72℃ final extension for 5 min. The PCR amplification program for the Lgc1 label is as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 50 s, 35 cycles; 72℃ final extension for 5 min.

[0035] In a preferred embodiment of the present invention, the cultivation method includes: S1: A hybrid F1 is obtained by crossing a rice variety with high dietary fiber as the female parent with a rice variety with aroma. S2: Plant F1, remove false hybrids, and harvest the seeds after maturity; S3: Plant F2, and use the aroma gene linkage marker (Badh-M2 molecular marker) and low gluten protein gene linkage marker (Lgc1 molecular marker) to detect the genotype of individual plants during the seedling stage, and screen out individual plants that are homozygous for both the aroma gene and the low gluten protein gene, and harvest individual plants for seed. S4: After soaking the seeds obtained in S3 in water, place them under a transmission optical detection device and screen for plump, opaque, chalky white seeds based on their light transmittance to determine them as high-dietary-fiber individuals. S5: Plant the seeds obtained from S4 screening, select the best single plants based on agronomic traits, and harvest seeds from a single plant with a single ear. The entire growth period of the superior single plant is 110-120 days, and its entire growth period is ≤ that of the rice variety Tianyou Huazhan; the number of effective tillers of the single plant is ≥8; S6: Plant the seeds obtained in S5, and repeat steps S4 and S5 to select seeds with good agronomic traits; S7: Seeds obtained from planting S6 are continuously self-pollinated until the phenotype is stable, and seeds are harvested from a single plant. S8: Seeds obtained from planting S7 were planted in plots and comparative trials were conducted. The gluten and dietary fiber content in the rice were tested. Strains with stable phenotypes, early growth period, high yield, low gluten content and high dietary fiber content were selected to complete the breeding process.

[0036] As a preferred embodiment, the present invention achieves the directional aggregation of aroma, low gluten protein and high dietary fiber traits through the complete breeding process of S1 to S8 described above. The resulting rice variety has the characteristics of rice aroma, low gluten protein, high dietary fiber and low glycemic index, and at the same time has good yield, early maturity, excellent plant and leaf morphology and wide ecological adaptability.

[0037] According to one aspect of the present invention, a low-gluten, high-dietary-fiber, and aromatic rice obtained by the above-described cultivation method is provided.

[0038] The low-gluten, high-dietary-fiber, high-fragrant rice provided by this invention has been tested and found to have the advantages of rice aroma, low gluten, high dietary fiber and low glycemic index. At the same time, it has good yield, early maturity, excellent plant and leaf morphology and wide ecological adaptability, making it a multifunctional and excellent rice.

[0039] In a preferred embodiment of the present invention, the rice has a gluten content of <4%, a dietary fiber content of ≥4.0%, and a rice aroma; The glycemic index (GI) of the rice is ≤35.

[0040] According to one aspect of the present invention, the use of the above-mentioned low-gluten, high-dietary-fiber, and aromatic rice in the preparation of functional staple food products; The functional staple food mentioned above is a functional staple food for patients with chronic kidney disease, diabetes, or obesity.

[0041] The low-gluten, high-fiber, and aromatic rice provided by this invention can be widely used in the preparation of functional staple food products, which are intended for patients with chronic kidney disease, diabetes, or obesity.

[0042] The technical solution of the present invention will be further described below with reference to the embodiments.

[0043] Example 1 Example 1 of this application uses Kangdian No. 1, a high dietary fiber variety, as the female parent and Jiugu No. 1, a fragrant low gluten protein variety, as the male parent for hybridization. The F2 generation seedlings were screened for double homozygous single plants using Badh-M2 and Lgc1 molecular markers, and high chalky seeds were screened by LED transmission after soaking for 3 hours. After multiple generations of agronomic trait selection and multi-point variety comparison tests, it was obtained and named "Jiugu No. 2".

[0044] Figure 1 This is a technical roadmap for the cultivation of the low-gluten, high-dietary-fiber aromatic rice variety "Jiugu No. 2" provided in this embodiment.

[0045] See Figure 1 The specific cultivation and verification methods are as follows: (I) Parental selection: Using Kangdian No. 1, a high-fiber indica rice variety (variety rights announcement number: CNA078407E), as the female parent, and Jiugu No. 1, a fragrant low-gluten indica rice variety (variety approval number: Gan Shen Dao 20220041; variety rights number: CNA20201000118), as the male parent, artificial hybridization was carried out at the Nanfan Experimental Base of Jiangxi Academy of Agricultural Sciences in Sanya City (109.4°E, 18.3°N) to obtain hybrid F1.

[0046] (II) Breeding process of multifunctional aromatic rice with low gluten protein and high dietary fiber: (1) Using Kangdian No. 1 as the female parent and Jiugu No. 1 as the male parent, F1 was obtained by hybridization; (2) Plant all F1s, remove false hybrids when they head and mature, and harvest at least 3,000 seeds after they mature. (3) 2040 F2 single plants were planted, and each single plant was tested by PCR electrophoresis using the aroma molecular marker Badh-M2 and the low gluten molecular marker Lgc1 during the seedling stage. Combined with agronomic trait selection, 44 single plants with homozygous aroma and low gluten genes were selected and harvested individually when mature. The above molecular marker detection methods are as follows: 1. Extraction of genomic DNA from rice plants; 2. Information on markers used in marker-assisted selection: The primer sequences for the Badh-M2 molecular marker are shown in SEQ ID NO. 1 and SEQ ID NO. 2; The sequence of SEQ ID NO. 1 is as follows: 5'-TAGGTTGCATTTACTGGGAG-3'; The sequence of SEQ ID NO. 2 is as follows: 5'-GGAAACAAACCTTAACCATAG-3'.

[0047] The Lgc1 molecular marker includes the Lgc1-1 primer sequence and the Lgc1-2 primer sequence; The Lgc1-1 primer sequences are shown in SEQ ID NO. 3 and SEQ ID NO. 4; The sequence of SEQ ID NO. 3 is as follows: 5'-TTCTACAATGAAGGCGATGC-3'; The sequence of SEQ ID NO. 4 is as follows: 5'-CTGGGCTTTAACGGGACT-3'; The Lgc1-1 primer sequences are shown in SEQ ID NO. 5 and SEQ ID NO. 6; The sequence of SEQ ID NO. 5 is as follows: 5'-ACCGTGTTATGGCAGTTT-3'; The sequence of SEQ ID NO. 6 is as follows: 5'-ATTCAAGGGCTATCGTCT-3'.

[0048] 3. PCR reaction system: 1 μL template DNA, 1 μL 10×PCR Buffer, 1 μL 2 mM dNTPs, 1 μL primer set, 0.2 μL Taq enzyme, add ddH2O to make up to 10 μL.

[0049] 4. PCR amplification conditions: The PCR amplification conditions for Badh-M2 labeling were: 94℃ pre-denaturation for 5 min, 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 20 s, 35 cycles of amplification, 72℃ extension for 5 min, and storage at 18℃. The PCR amplification conditions for Lgc1 labeling were: 94℃ pre-denaturation for 5 min, 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 50 s, 35 cycles of amplification, 72℃ extension for 5 min, and storage at 18℃. 5. Electrophoresis detection of amplification products: PCR products of Badh-M2 were detected by 6% non-denaturing polyacrylamide gel electrophoresis; The PCR products of Lgc1 were subjected to 2.5% agarose gel electrophoresis.

[0050] Figure 2 This is a graph showing the electrophoresis results of the F2 generation Badh-M2 molecular marker PCR amplification provided in this embodiment; Depend on Figure 2 It can be seen that lanes 1-63 are the results of Badh-M2 molecular marker electrophoresis for F2 generation rice seedlings to be screened. Among them, the single plants with the same banding pattern as Jiugu No. 1 carry the homozygous aroma gene.

[0051] Figure 3 This is a graph showing the electrophoresis results of Lgc1 molecular marker PCR amplification in the F2 generation population provided in this embodiment. Depend on Figure 3 It can be seen that lanes 1-20 are the electrophoresis results of Lgc1 molecular markers for F2 generation rice to be screened. Among them, the single plants with the same banding pattern as Jiugu No. 1 carry the homozygous low glutenin gene.

[0052] (4) After the harvested seeds with double homozygous genes of aroma and low glutenin are dried, they are soaked in water for 3 hours. After soaking, the seeds are placed on a high-brightness LED projection lamp table. The plump, opaque, highly chalky seeds containing high dietary fiber are selected by passing the light through them. A total of 13 samples were selected and then dried. (5) Plant the 13 selected seeds according to the lineage. Plant all seedlings of each lineage and select 6 superior single plants with a growth period earlier than Tianyou Huazhan, medium or above tillering ability and good plant and leaf morphology. Harvest seeds from single ears. (6) Plant F4, repeat steps 4 and 5, select the best from the best, and retain 3 superior strains; (7) Plant F5 and self-pollinate continuously until the phenotype is basically stable in F6. Plant at least 48 plants per generation for each line and harvest seeds from each plant individually. (8) Each of the three harvested individual plants was planted in a plot of 300 plants. The overall performance and yield were assessed, and the gluten and dietary fiber content in the rice was tested. Through comprehensive comparison, a strain with stable phenotype, early maturity, high yield, low gluten content and high dietary fiber content was selected and named Jiugu No. 2. Its rice has a gluten content of 2.8%, which is less than one-third of that of ordinary rice, and a dietary fiber content of 4.22%, which is 8 times that of ordinary rice. Its GI value is 32.7, which is a low GI food.

[0053] Experimental Example 1 To verify the technical effectiveness of the cultivation method of the present invention, the rice obtained in Example 1 was simultaneously tested in five representative rice ecological zones in Jiangxi Province in 2023: Nanchang, Fengcheng, Gao'an, Zhangshu, and Anfu. Each plot was 0.5 mu in size, with Tianyou Huazhan as the control.

[0054] (II) Testing Methods: (1) The method for detecting glutenin is as follows: Rice seeds were dehulled into brown rice and ground into powder using a mortar and pestle. 0.05 g of rice powder was weighed and transferred to a 1.5 mL centrifuge tube. 1 mL of protein extraction buffer was added. The extraction buffer consisted of 0.25 M Tris-HCl (pH 6.8), 8 M urea, 4% SDS, 5% β-mercaptoethanol, and 20% glycerol. The mixture was vortexed thoroughly for several seconds and then placed in a shaker at 25°C for 12 h (overnight). The mixture was centrifuged at 10000 r / min for 10 min, and the supernatant was transferred to a 5 mL centrifuge tube. This centrifugation process was repeated twice. 10 μL of the supernatant was used for SDS-PAGE electrophoresis (15%). After electrophoresis, the gel was soaked in fixative (10% acetic acid, 40% ethanol, 50% distilled water) for 2 h. The gel was then washed 3-4 times with water for 15 min each time and stained with Coomassie Brilliant Blue (overnight). Dye solution formula: 0.12g Coomassie Brilliant Blue G250, 10g ammonium sulfate, 10mL phosphoric acid, 20mL methanol, 70mL distilled water. Decolorize for at least 5 hours (shaking constantly in distilled water).

[0055] (2) The method for detecting dietary fiber is in accordance with GB 5009.88-2023 National Food Safety Standard - Determination of Dietary Fiber in Food.

[0056] (3) GI value determination: The in vitro simulated digestion method was used, with glucose as a reference, to calculate the hydrolysis rate and convert the GI value. For details, refer to BJC-FDD-FB2105 "Simulated Digestion Analysis of Glycemic Index of Food".

[0057] (4) Multi-point ecological adaptability test: In 2023, it was carried out simultaneously in five representative rice ecological zones in Nanchang, Gao'an, Zhangshu, Fengcheng and Anfu in Jiangxi Province. The plot area was 0.5 mu, and Tianyou Huazhan was used as the control to compare yields.

[0058] (III) The experimental results are as follows: 1. Agronomic traits and yield performance at each site, as well as gluten content, dietary fiber content, and GI value (2023 Jiangxi five-site joint experiment), are detailed in Table 1.

[0059] Table 1. Experimental results of rice (Jiugu No. 2) in Example 1 of this application:

[0060] As shown in Table 1, the growth period of the rice (Jiugu No. 2) in Example 1 of this application is not longer than that of Tianyou Huazhan in various regions, the yield of dry grains per mu is more than 450 kg, the gluten content is 2.7~3.1%, the dietary fiber content is 4.1~4.8%, and the GI value is 31.3~38.2.

[0061] To visually present the agronomic phenotype, appearance quality, and proteomics characteristics of the rice variety "Jiugu No. 2" in Example 1 of this invention, the following image data were simultaneously collected and analyzed. All images are from the stable "Jiugu No. 2" lines cultivated in Example 1, and the shooting / electrophoresis conditions were strictly standardized: Figure 4 The field phenotype of a typical single plant of Kutani No. 2 in Example 1 provided for this experimental case. Figure 4 It can be seen that the Jiugu No. 2 plant has a moderate height (about 104 cm) and plant shape, upright sword leaves, and strong tillering ability.

[0062] Figure 5 The image shows the appearance of Kutani No. 2 polished rice provided for this experimental example. (Source: [Original Source Name]) Figure 5 It can be seen that Jiugu No. 2 rice has a high chalkiness rate.

[0063] Figure 6 This is the SDS-PAGE electrophoresis pattern of total protein from Jiugu No. 2 rice provided for this experimental example. Figure 6 It can be seen that the gluten content of Jiugu No. 2 is similar to that of Jiugu No. 1, and significantly lower than that of Kangdian No. 1.

[0064] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for cultivating a low-gluten, high-dietary-fiber, high-aroma rice variety, characterized in that, The cultivation method includes: By using hybridization, rice varieties with high dietary fiber were used as the female parent to cross with fragrant rice varieties. The hybrid offspring were then subjected to marker-assisted selection using DNA molecular markers, combined with agronomic trait selection, to obtain high-quality fragrant rice with low gluten content and high dietary fiber. The marker-assisted selection begins in the F2 generation, and the markers include Badh-M2 and Lgc1 markers.

2. The cultivation method according to claim 1, characterized in that, The molecular marker-assisted selection includes: During the F2 generation rice seedling stage, the genotype of individual plants was detected using Badh-M2 and Lgc1 molecular markers to screen out individual plants that were homozygous for both the Badh2 aroma gene and the Lgc1 low glutenin gene. Then, the seeds of the planted individual plants were soaked in water and placed under a transmission optical detection device. Based on the light transmittance of the seeds, plump, opaque, chalky seeds were selected.

3. The cultivation method according to claim 2, characterized in that, The seeds are soaked in water for 3 to 5 hours, preferably 3 hours.

4. The cultivation method according to claim 1, characterized in that, The primer sequences for the Badh-M2 molecular marker are shown in SEQ ID NO. 1 and SEQ ID NO. 2; The sequence of SEQ ID NO. 1 is as follows: 5'-TAGGTTGCATTTACTGGGAG-3'; The sequence of SEQ ID NO. 2 is as follows: 5'-GGAAACAAACCTTAACCATAG-3'.

5. The cultivation method according to claim 1, characterized in that, The Lgc1 molecular marker includes the Lgc1-1 primer sequence and the Lgc1-2 primer sequence; The Lgc1-1 primer sequences are shown in SEQ ID NO. 3 and SEQ ID NO. 4; The sequence of SEQ ID NO. 3 is as follows: 5'-TTCTACAATGAAGGCGATGC-3'; The sequence of SEQ ID NO. 4 is as follows: 5'-CTGGGCTTTAACGGGACT-3'; The Lgc1-1 primer sequences are shown in SEQ ID NO. 5 and SEQ ID NO. 6; The sequence of SEQ ID NO. 5 is as follows: 5'-ACCGTGTTATGGCAGTTT-3'; The sequence of SEQ ID NO. 6 is as follows: 5'-ATTCAAGGGCTATCGTCT-3'.

6. The cultivation method according to claim 1, characterized in that, The reaction system for the marker-assisted PCR amplification is 10 μL, comprising: 1 μL template DNA, 1 μL 10×PCR Buffer, 1 μL 2 mmol / L dNTP, 1 μL primer combination, 0.2 μL Taq DNA polymerase, and the remainder is sterile deionized water. The PCR amplification program for the Badh-M2 molecular marker was as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 20 s, 35 cycles; 72℃ final extension for 5 min. The PCR amplification program for the Lgc1 molecular marker was as follows: 94℃ pre-denaturation for 5 min; 94℃ denaturation for 20 s, 55℃ annealing for 20 s, 72℃ extension for 50 s, 35 cycles; 72℃ final extension for 5 min.

7. The cultivation method according to claim 1, characterized in that, The cultivation method includes: S1: A hybrid F1 is obtained by crossing a rice variety with high dietary fiber as the female parent with a rice variety with aroma. S2: Plant F1, remove false hybrids, and harvest the seeds after maturity; S3: Plant F2, and use aroma gene linkage markers and low gluten protein gene linkage markers to perform genotyping on individual plants during the seedling stage, and screen out individual plants that are homozygous for both aroma gene and low gluten protein gene, and harvest individual plants for seed. S4: After soaking the seeds obtained in S3 in water, place them under a transmission optical detection device and screen for plump, opaque, chalky white seeds based on their light transmittance to determine them as high-dietary-fiber individuals. S5: Plant the seeds obtained from S4 screening, select the best single plants based on agronomic traits, and harvest seeds from a single plant with a single ear. The entire growth period of the superior single plant is 110-120 days, and its entire growth period is ≤ that of the rice variety Tianyou Huazhan; the number of effective tillers of the single plant is ≥8; S6: Plant the seeds obtained from S5, and repeat steps S4 and S5 to select individual plants with good agronomic traits. S7: Seeds obtained from planting S6 are continuously self-pollinated until the phenotype is stable, and seeds are harvested from a single plant. S8: Seeds obtained from S7 were planted in plots and comparative trials were conducted. At the same time, the gluten and dietary fiber content in the rice were tested. Through comparison, strains with stable phenotypes, early growth period, high yield, low gluten content and high dietary fiber content were selected to complete the breeding.

8. The low-gluten, high-dietary-fiber, high-fragrant rice cultivated by the cultivation method according to any one of claims 1 to 7.

9. The low-gluten, high-dietary-fiber, high-aroma rice according to claim 8, characterized in that, Rice has a gluten content of <4%, a dietary fiber content of ≥4.0%, and a rice aroma; The glycemic index (GI) of rice is ≤55.

10. The use of the low-gluten, high-fiber, and aromatic rice according to claim 8 or 9 in the preparation of functional staple food products; The functional staple food mentioned above is a functional staple food for patients with chronic kidney disease, diabetes, or obesity.

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