Wxb (CT) 18 and SSIIaHX3 allele combination and application thereof in rice cooking taste quality improvement

Abstract

The invention discloses a novel Wx and SSIIa allele combination Wxb (CT) 18 and SSIIaHX3. The Wxb (CT) 18 is characterized in that a 5 '-UTR region of a Wx gene has 18 CT continuous repetitive sequences; the SSIIaHX3 is characterized in that the 264th site of the first exon of the SSIIa gene has Cgt; g, single nucleotide variation. The Wxb (CT) 18 and SSIIaHX3 specific allele combination can reduce the amylose content of rice, increase the medium-long-chain starch content, the gel consistency and the gelatinization temperature and improve the cooking taste quality of the rice, can efficiently improve the cooking taste quality of the rice in combination with molecular marker-assisted screening, and has an important value in rice variety breeding.

Description

Technical Field

[0001] The present invention relates to the technical field of rice breeding, and in particular to a Wx b (CT) 18 and SSIIa HX3 Allele combinations and their application in improving rice cooking and eating quality. Background Art

[0002] Rice ( Oryza sativa As one of the world's most important food crops, the cooking and flavor quality of rice directly influences consumer choice and market demand. As living standards improve, people's expectations for rice's flavor quality are becoming increasingly higher. The cooking and flavor quality of rice is primarily determined by physical and chemical parameters such as amylose content, gelatinization consistency, and gelatinization temperature. Wx Genes and SSIIa These genes are two important main effect genes that affect the cooking and eating quality of rice, regulating the amylose content and gelatinization temperature respectively.

[0003] Currently, studies have shown that Wx Different alleles of a gene (e.g. Wx a 、 Wx b 、 wx etc.) have a significant effect on the amylose content of rice, while SSIIa Different alleles of the gene mainly affect the gelatinization temperature of rice. Wx and SSIIa There are some reports on the effects of gene combinations on rice quality, such as Wx mp genetic background SSIIa and PUL The article "Genetic Effects of Cooking and Taste Quality Traits" points out that Wx mp Discussion under genetic background SSIIa and PUL Genetic effects of genes on cooking and eating quality traits from Wuyunjing No. 21 SSIIa b Genes and PUL b The genes increased the amylose content by 0.29% to 1.00% and 0.62% to 1.18%, respectively, and PULThe effect of SSIIa is greater than that of SSIIa, and there is an interaction effect between the two. Patent CN117965701A discloses that the genotypes of three KASP markers Wx-In1-KASP, Wx-Ex10-KASP, and SSIIa-KASP of rice are detected, and the cooking and eating quality of the tested rice is determined according to the genotype. Therefore, further exploration of new Wx and SSIIa The effects of allele combinations on rice quality can provide an important theoretical basis for improving and breeding high-quality rice materials. Summary of the Invention

[0004] The purpose of the present invention is to overcome the above-mentioned defects and deficiencies in the prior art and to provide a Wx b (CT) 18 and SSIIa HX3 Allele combination, the Wx b (CT) 18 for Wx The 5'-UTR region of the gene has 18 consecutive CT repeat sequences; SSIIa HX3 for SSIIa There is a C>G single nucleotide variation at position 264 of the first exon of the gene.

[0005] The second object of the present invention is to provide the above Wx b (CT) 18 and SSIIa HX3 Allele combination and including the above Wx b (CT) 18 and SSIIa HX3 Application of biological materials with allele combinations in improving or testing the cooking and eating quality of rice.

[0006] The third object of the present invention is to provide a method for improving the cooking and eating quality of rice.

[0007] A fourth object of the present invention is to provide a method for detecting the cooking and eating quality of rice.

[0008] A fifth object of the present invention is to provide a method for detecting the above Wx b (CT) 18 and SSIIa HX3 Application of primer sets of allele combinations in assisted selection breeding for improving rice cooking and eating quality.

[0009] The above-mentioned object of the present invention is achieved through the following technical solutions: The present invention provides a Wx b (CT) 18 and SSIIa HX3 Allele combination, the Wx b (CT) 18 for Wx The 5'-UTR region of the gene has 18 consecutive CT repeat sequences; SSIIa HX3 for SSIIa There is a C>G single nucleotide variation at position 264 of the first exon of the gene.

[0010] The present invention discovered a special rice germplasm Hangxiang Si Miao No. 3 Wx b (CT) 18 and SSIIa HX3 The combination of specific alleles can reduce the amylose content of rice, increase the medium and long chain starch content, gel consistency and gelatinization temperature, and improve the cooking and eating quality of rice. Wx and ​ The gene sequence was amplified and sequenced, and it was found that ​ Gene ​ b (CT) 18 type, and ​ The single nucleotide variant at the Ex1-264 C>G position is a missense mutation, resulting in an amino acid change from aspartic acid to glutamic acid, so this allele is named ​ HX3 In order to explore the factors that affect the cooking and eating quality of Hangxiang Si Miao No. 3, the present invention screened several different rice varieties and ​ and ​ Genotype classification was performed and it was found that ​ b (CT) 18 and ​ HX3 The rice varieties all have excellent cooking and eating quality. ​ b (CT) 18 ​ HX3 The fragments were introduced into Huahang Special Rice, and continuous backcrossing was used to construct a near-isogenic line. The rice amylose content of this combination was reduced, the gel consistency was increased, and the taste was significantly improved, proving that this combination can significantly improve the cooking and taste quality of the recipient material.​ b (CT) 18 and ​ HX3 The rice materials of the genotype combination have the following characteristics: the amylose content is 13%-18% and the gel consistency is above 70mm. ​ b (CT) 18 ​ HX3 It's a new ​ and ​ Allele combination, which can significantly improve the cooking and eating quality of rice, is of great significance in controlling the cooking and eating quality of rice.

[0011] Furthermore, the ​ b (CT) 18 The genome sequence is shown in SEQ ID NO.1 or SEQ ID NO.2; ​ HX3 The genome sequence is shown in SEQ ID NO.4.

[0012] The present invention also provides the above ​ b (CT) 18 and ​ HX3 Application of allele combinations in improving or detecting rice cooking and eating quality.

[0013] The present invention also provides the above ​ b (CT) 18 and ​ HX3 Application of biological materials with allele combinations in improving or detecting the cooking and eating quality of rice; the biological materials are expression cassettes, expression vectors, and engineered bacteria.

[0014] Furthermore, the improved rice cooking and eating quality is to reduce the amylose content of rice and increase the medium and long chain starch content, gel consistency and gelatinization temperature.

[0015] Furthermore, the improvement is to control the amylose content to 13% to 18% and the gel consistency to be above 70mm.

[0016] Preferably, the rice is silk rice, which improves the cooking quality of silk rice.

[0017] The present invention also provides a method for improving the cooking and eating quality of rice, wherein the method comprises the following steps: ​ and ​ Gene replacement as above​ b (CT) 18 and ​ HX3 Allele combination.

[0018] Furthermore, the replacement method is to replace the ​ b (CT) 18 and ​ HX3 The present invention constructs different ​ b (CT) 18 and ​ HX3 The effect of the combined gene was verified by using a near-isogenic line with a combination of alleles. It was found that the near-isogenic line had a lower amylose content, higher gel consistency, and significantly improved taste compared to the recipient. ​ and ​ The new method of improving the cooking and eating quality of rice by gene combination has important breeding application value.

[0019] The present invention also provides a method for detecting the cooking and eating quality of rice, wherein the method is to detect the ​ and ​ The genotype of the gene, when the above ​ b (CT) 18 and ​ HX3 The allele combination includes rice with low amylose content, increased medium- and long-chain amylopectin content, gel consistency and gelatinization temperature, and better rice cooking and eating quality.

[0020] Furthermore, the detection of the rice to be tested ​ and ​ The primer sequences for the genotype of the gene are shown in SEQ ID NOs. 8-9 and SEQ ID NOs. 14-15.

[0021] The present invention also provides a method for detecting the above ​ b (CT) 18 and ​ HX3 The primer set of the allele combination is used in assisted selection breeding for improving rice cooking and eating quality. The primer set is shown in SEQ ID NOs. 8-9 and SEQ ID NOs. 14-15.

[0022] Compared with the prior art, the present invention has the following beneficial effects: The present invention provides a new ​ and ​ Allele combination ​ b (CT) 18 and ​ HX3 , ​ b (CT) 18 For ​ The CT sequence in the 5'-UTR region of the gene is repeated 18 times; ​ HX3 For ​ The 264th single nucleotide mutation of the first exon of the gene is G. ​ b (CT) 18 and ​ HX3 The specific allele combination can reduce the amylose content of rice, increase the medium and long-chain starch content, gel consistency and gelatinization temperature, and improve the cooking and taste quality of rice. Combined with molecular marker-assisted screening, it can effectively improve the cooking and taste quality of rice, which is of great value in rice variety breeding. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] ​ for ​ Schematic diagram of gene segmentation sequencing.

[0024] ​ Hangxiang Silk Seedling No. 3 ​ Gene sequence comparison results.

[0025] ​ for ​ Schematic diagram of gene segmentation sequencing.

[0026] ​ Hangxiang Silk Seedling No. 3 ​ Gene sequence comparison results.

[0027] ​ Comparative analysis of cooking and eating quality traits of different allele combinations. Note: ANOVA test, significant markers based on Duncan test results.

[0028] ​ Comparative analysis of thermal characteristics related to different allele combinations. Note: To: starting temperature, Tp: peak temperature, Tc: ending temperature; ANOVA test; significance marks based on Duncan's test results.

[0029] ​ The distribution of amylopectin chain lengths for different allele combinations. Note: ANOVA test, significance marks based on Duncan's test results.

[0030] ​ Special TZ for China Airlines ​ b ​ HX3 Near-isogenic lines.

[0031] ​ This paper analyzes the cooking and eating quality traits of near-isogenic lines.

[0032] ​ This is the RVA spectrum curve analysis of the near-isogenic line. DETAILED DESCRIPTION

[0033] The present invention will be further described below with reference to the accompanying drawings and specific examples, but the examples do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0034] Unless otherwise specified, the reagents and materials used in the following examples were commercially available.

[0035] Example 1 Hangxiang Silk Seedling No. 3 ​ and ​ Gene sequence analysis Hangxiang Si Miao No. 3 is a rice variety selected and bred by the National Plant Space Breeding Engineering Technology Research Center of South China Agricultural University and is preserved in the center.

[0036] 1. ​ Gene sequencing right ​ The gene sequence amplification is divided into three fragments, labeled W1, W2 and W3, and the product sizes are 1.8 kb, 1.8 kb and 1.8 kb respectively. ​The primer sequences for the three amplified fragments are shown in Table 1. PCR amplification was performed using a 30 μL system consisting of the following components: 15 μL 2× PCR Taq Master Mix (containing Taq DNA Polymerase, dNTPs, PCR buffer, and MgCl₂), 1 μL Forward Primer, 1 μL Reverse Primer, 1 μL DNA (100 ng / mL), and 12 μL ddH₂O. The PCR reaction program was as follows: initial denaturation at 95°C for 3 min; denaturation at 95°C for 15 s; annealing at 58°C for 30 s; and extension at 72°C for 2 min. This cycle of denaturation, annealing, and extension was repeated for 35 cycles, followed by a final extension at 72°C for 5 min and a 20°C incubation period. After the reaction, the DNA fragments were stored at 4°C until further use. Sequencing of the amplified DNA fragments was performed by the Guangzhou Branch of Beijing Qingke Biotechnology Co., Ltd. Sequence alignment of all the determined gene sequences was performed using MEGA7 software.

[0037] Table 1 For amplification ​ Primers and sequences of gene fragments

[0038] By overlapping and splicing the sequences of three sequencing fragments, we obtained ​ The complete gene sequence is 5032 bp, and the nucleotide sequence is shown in SEQ ID No. 1. ​ Sequence analysis results are as follows ​ As shown, it shows that Hangxiang Si Miao No. 3 ​ Genes are ​ b (CT) 18 type.

[0039] 2. ​ Gene sequencing right ​ The sequence amplification of the gene was divided into five fragments, labeled S2-1, S2-2, S2-3, S2-4 and S2-5, and the product sizes were 1.4 kb, 1.5 kb, 1.4 kb, 1.4 kb and 0.7 kb, respectively. ​The primer sequence information for the five amplified fragments is shown in Table 2. The PCR amplification system was the same as above. The PCR reaction program was as follows: pre-denaturation at 95°C for 3 min; denaturation at 95°C for 15 s; annealing at 49–51°C for 30 s; extension at 72°C for 2 min; denaturation, annealing, and extension for 35 cycles; final extension at 72°C for 5 min; and insulation at 20°C. After the reaction, the sample was stored at 4°C for future use. Sequencing of the amplified DNA fragments was completed by the Guangzhou Branch of Beijing Qingke Biotechnology Co., Ltd. Sequence alignment was performed using MEGA7 software based on all the gene sequences measured.

[0040] Table 2 For amplification ​ Primers and sequences of gene fragments

[0041] By overlapping and splicing the sequences of 5 sequencing fragments, we obtained ​ The complete gene sequence is 4905 bp, and the nucleotide sequence is shown in SEQ ID No.4. ​ Sequence analysis results are as follows ​ The results showed that EX1-264C>G of Hangxiang Si Miao 3 was a new allele variation, and the allele type of Hangxiang Si Miao 3 was named ​ HX3 .

[0042] Based on the above results, the ​ b (CT) 18 and ​ HX3 Allele combination.

[0043] Example 2 Analysis of the cooking and eating quality of Hangxiang Si Miao No. 3 1. Experimental Methods For analysis ​ b (CT) 18 and ​ HX3 The effects of specific combinations on the cooking and eating quality of Hangxiang Si Miao 3 were studied. The silk rice varieties approved by Guangdong Province, including Xiangyaxiangzhan, Meixiangzhan 2, Yuzhenxiang, Hangjuxiang Si Miao, 19xiang, Zengkexinxuan Si Miao 1, Huahangxiang Yinzhen, Huahang 51 and Huahang 57, were selected as controls. H273, H274 and H275 are carriers obtained by hybridization of Hangxiang Si Miao 3 and Hangjuxiang Si Miao. ​ b (CT)18 and ​ HX3 The special combination high-generation silk seedling rice line is preserved in the National Plant Space Breeding Engineering Technology Research Center of South China Agricultural University and has the same characteristics as Hangxiang Silk Seedling No. 3. ​Allele type ​ HX3 China Airlines Flight 51 and China Airlines Flight 57 ​ The 5' end splicing site upstream of the leader intron of the gene carries 17 CT repeat sequences. ​ b (CT) 17 Allele type, the rest of the breeds carry the same ​ b (CT) 18 Allele types are shown in Tables 3 and 4.

[0044] Hangxiang Silk Seedling No. 3, H273, H274, H275, 19 incense, Huahang incense silver needle, Zengke No. 1, Hangjuxiang Silk Seedling ​ b (CT) 18 The sequence is shown as SEQ ID No.1.

[0045] Ivory incense, beautiful incense, jade needle incense ​ b (CT) 18 The sequence is shown as SEQ ID No.2.

[0046] China Airlines Flight 51 and 57 ​ b (CT) 17 The sequence is shown as SEQ ID No.3.

[0047] Hangxiang Silk Seedling No. 3, H273, H274, H275 ​ HX3 The sequence is shown as SEQ ID No.4.

[0048] 19 incense, Zengke No. 1, China Airlines silver needle, ivory incense, beauty incense, jade needle incense, China Airlines 51, China Airlines 57, Hangju incense silk seedling ​ b The sequence is shown as SEQ ID No.5.

[0049] Table 3 Different varieties ​ Differences in gene mutation sites

[0050] Table 4 Different varieties ​ Differences in gene mutation sites

[0051] The rice quality of all the above varieties was tested, including amylose content, gelatin consistency, alkali elimination value, gelatinization temperature and amylopectin structure. The specific experimental methods are as follows: (1) Determination of amylose content: Weigh 10 mg ± 0.1 mg of rice flour (standard samples with AAC of 1.1%, 9.6%, 15.9%, and 26.1%) and the test sample into a 25 mL volumetric flask, with three replicates for each standard sample and test sample. Add 250 μL of 95% ethanol solution to the volumetric flask, shake the volumetric flask gently to moisten and disperse the sample, and then add 2.25 mL of 1 mol / L NaOH solution along the neck of the volumetric flask. Then, place the volumetric flask in a 100°C water bath and boil for 10 min. Take it out and wait for the boiled sample to cool to room temperature, then dilute to volume and shake well. 1.25 mL of the test sample solution was pipetted into a new 25 mL volumetric flask. 250 μL of 1 mol / L acetic acid solution and 375 μL of iodine-potassium iodide solution were added to the new flask containing the test sample solution, the volume was adjusted, the mixture was shaken, and the reaction was allowed to react for 15-20 min. 1.25 mL of 0.09 mol / L sodium hydroxide solution was added to the new 25 mL volumetric flask instead of the test sample solution. Acetic acid solution and iodine-potassium iodide solution were added in the same manner to prepare a test blank solution. Using a visible spectrophotometer (7230G, Shanghai Youke Instrument Co., Ltd., China), the absorbance of each test sample was measured at a wavelength of 620 nm, with the prepared blank solution set to zero. The AAC of the standard sample and the corresponding absorbance were used to construct a standard curve, and the AAC of the test sample was calculated according to the obtained regression equation.

[0052] (2) Determination of gel consistency: Accurately weigh 100 mg ± 1 mg of the rice flour sample to be tested into a test tube; add 0.2 mL of 0.025% bromothymol blue ethanol solution (weigh 125 mg of thymol blue dissolved in 500 mL of 95% ethanol), and use a vortex mixer to fully disperse the sample; then add 2.0 mL of 0.200 mol / L KOH solution, use a vortex mixer to fully disperse the sample, immediately place the test tube in a boiling water bath and start the timer, cover the test tube mouth with glass beads, and heat for 8 minutes; after heating is completed, take out the test tube and remove the lid, let it stand at room temperature for 5 minutes, and then put the test tube into ice prepared in advance to cool for 20 minutes; take the test tube out of the ice and immediately place it horizontally on a scaled horizontal surface, aligning the bottom of the test tube with the starting scale line, and let it stand at 25 ± 2 ° C for 1 hour, and measure the length from the bottom of the test tube to the front end of the rice glue flow in the test tube in millimeters (mm).

[0053] (3) Alkali digestion value determination: Randomly select 6 whole and full grains of polished rice from the whole polished rice to be tested and place them in a 90 mm diameter culture dish with 3 replicates. Add 20.0 mL of 1.7% (m / v) KOH solution and immediately use a glass rod to evenly distribute the rice grains in the dish. Cover the dish and incubate at a constant temperature of 30 ± 2 °C for 23 h. Then, observe the digestion of the endosperm of each grain and record the results according to the standard.

[0054] (4) Determination of gelatinization temperature: The thermal properties of the samples were measured using a DSC8000 (PerkinElmer Medical Diagnostics Shanghai Co., Ltd., USA). The samples were mixed with water with a moisture content of approximately 70% and equilibrated at 25°C for 12 h. Then, the temperature was scanned from 25°C to 125°C at a heating rate of 10°C / min. Thermal parameters such as the onset temperature (To), peak temperature (Tp), end temperature (Tc), and transition enthalpy (ΔH) were calculated from the DSC endothermic curve.

[0055] (5) Determination of amylopectin structure: The separated starch was debranched with isoamylase and then subjected to starch fine structure analysis. Amylopectin chain length distribution (CLD) was analyzed using a high performance anion exchange chromatography (HPAEC) system on a CarboPac PA-200 anion exchange column (4.0 × 250 mm; Dionex) and a pulsed amperometric detector (PAD). Amylopectin chain length distribution (CLD) was analyzed on a Dionex ICS 5000 system.

[0056] 2. Experimental Results The results are as follows ​ As shown, carrying ​ b (CT) 18 ​ HX3 Rice varieties with specific allele combinations are compared with those carrying ​ b (CT) 18 ​ b 、 ​ b (CT) 17 ​ b Rice varieties with specific allele combinations have significantly reduced amylose content and alkali digestibility, and significantly increased gel consistency; ​ b (CT) 17 ​ b Rice varieties with specific allele combinations have higher amylose content and a harder texture. ​ b (CT) 18 ​HX3 The specific allele combination plays an important role in improving the soft and elastic taste of rice, and can make rice have better steaming and cooking taste quality.

[0057] like ​ As shown, carrying ​ b (CT) 18 ​ HX3 The rice varieties with the specific allele combination had a significantly broader endothermic peak, while the other two allele combinations had narrower endothermic peaks. The onset temperature (To), peak temperature (Tp), and termination temperature (Tc) of the three combinations all showed significant differences. ​ b (CT) 18 ​ HX3 The specific allele combination had higher starting temperature, peak temperature and ending temperature, indicating that the variety carrying this allele combination had a higher gelatinization temperature.

[0058] like ​ As shown, ​ b (CT) 18 ​ HX3 The content of short chain (A chain) of the specific allele combination is significantly lower than that of the other two combinations, while the content of medium and long chain amylopectin is higher than that of the other two combinations, especially the B1 chain. The taste is soft and elastic, and the rice has excellent cooking and eating quality (more medium and long chains indicate that the starch of this variety is slowly digested and has a high gelatinization temperature, which is consistent with the results measured by DSC). ​ b (CT) 18 ​ HX3 The gelatinization temperature of the combination is high).

[0059] Example 3 ​ b (CT) 18 ​ HX3 Verification of the effects of specific allele combinations on cooking and eating quality Huahang Tezhan (TZ) is a high-amylose indica rice variety bred by South China Agricultural University. ​ a ​ b Genotype combination (sequences are shown in SEQ ID NO.6 and 7). ​ b (CT) 18 ​ HX3The fragment was introduced into the China Airlines special account, and after four generations of backcrossing and two generations of self-crossing, three BC4F3 generations of TZ were finally obtained. - ​ b ​ HX3 Near isogenic lines, NIL-1, NIL-2 and NIL-3, have plant shapes like ​ shown.

[0060] The cooking and eating quality traits of the recipient parent TZ and the donor parent Hangxiang Si Miao No. 3 (HX3) were measured and analyzed: (1) The determination methods of amylose, gel consistency and alkali elimination value are as in Example 2 above.

[0061] (2) Determination of RVA spectrum characteristic values: The viscosity characteristics of rice starch were determined using an RVA instrument (Rapid Visco Analyser-TecMaster, Perten, Sweden), and the supporting software TCW 3 (ThermoCline for Windows) was used to collect and organize the data. The RVA spectrum characteristic values ​​of rice were determined according to the standard method of the American Cereal Chemists Association. The specific method is as follows: when the moisture content of rice flour is 12%, 3.00 g of sample rice flour is weighed and placed in a measuring tank. 25.00 mL of distilled water is added and a paddle is installed. The temperature in the tank is changed according to a predetermined program: 50°C for 1 min, then 12°C min -1 The temperature in the tank was raised to 95°C (3.75 min) at an increasing rate, maintained at 95°C for 2 min, then dropped to 50°C at the same rate (3.75 min), and maintained at 50°C for 1.4 min. The fixed-blade agitator rotated at a rate of 960 r·min in the initial 10 s. -1 , and then maintain at 160 r / min.

[0062] The results are as follows ​ As shown, the recipient parent TZ has the highest amylose content, up to 23.3%, while the donor parent HX3 has an amylose content of 9.3%. The amylose content of the three near-isogenic lines is significantly different from that of their parents, at 15.9%, 18.6%, and 16.6%, respectively. The recipient parent TZ has the lowest gel consistency, at only 22.3 mm, while the donor parent HX3 has a gel consistency of 81.7 mm. There is no significant difference between the gel consistency of the three near-isogenic lines and that of HX3. The results of the alkali dissolution value analysis show that the recipient parent TZ has the highest alkali dissolution value, which is significantly higher than that of the donor parent and the three near-isogenic lines. ​ As shown in Figure 2, the RVA profiles of the three near-isogenic lines are similar to those of the donor parent HX3, with a higher peak viscosity and a lower cold gel viscosity than that of the recipient parent TZ.​ b (CT) 18 ​ HX3 The rice with the specific allele combination has a soft texture and excellent cooking and eating quality.

Claims

1. A Wx b (CT) 18 and SSIIa HX3 An allele combination characterized in that described Wx b (CT) 18 for Wx The 5'-UTR region of the gene has 18 consecutive CT repeat sequences; SSIIa HX for SSIIa There is a C>G single nucleotide variation at position 264 of the first exon of the gene.

2. according to claim 1 Wx b (CT) 18 and SSIIa HX3 An allele combination characterized in that described Wx b (CT) 18 The genome sequence is shown in SEQ ID NO.1 or SEQ ID NO.2; SSIIa HX3 The genome sequence is shown in SEQ ID NO.

4.

3. The method according to claim 1 or 2 Wx b (CT) 18 and SSIIa HX3 Application of allele combinations in improving or detecting rice cooking and eating quality.

4. Containing the method according to claim 1 or 2 Wx b (CT) 18 and SSIIa HX3 Application of biological materials with allele combinations in improving or detecting the cooking and eating quality of rice; the biological materials are expression cassettes, expression vectors, and engineered bacteria.

5. The use according to claim 3 or 4, characterized in that: The improved rice cooking and eating quality comprises reducing the amylose content of rice and increasing the medium- and long-chain starch content, gel consistency and gelatinization temperature.

6. A method for improving the cooking and eating quality of rice, characterized in that: The method is to Wx and SSIIa Gene replacement as described in claim 1 or 2 Wx b (CT) 18 and SSIIa HX3 Allele combination.

7. The method according to claim 6, characterized in that The replacement method is to replace by genetic engineering means or by hybridization-backcrossing method. Wx b (CT) 18 and SSIIa HX3 introduced into recipient rice plants.

8. A method for detecting the cooking and eating quality of rice, characterized in that: The method is to detect the Wx and SSIIa The genotype of the gene as claimed in claim 1 or 2, when detected in rice Wx b (CT) 18 and SSIIa HX3 The allele combination includes rice with low amylose content, increased medium- and long-chain amylopectin content, gel consistency and gelatinization temperature, and better rice cooking and eating quality.

9. The method according to claim 8, characterized in that The method for detecting the rice to be tested Wx and SSIIa The primer sequences for the genotype of the gene are shown in SEQ ID NOs. 8-9 and SEQ ID NOs. 14-15.

10. Detection of claim 1 or 2 Wx b (CT) 18 and SSIIa HX3 The application of a primer set of allele combination in assisted selection breeding for improving rice cooking and eating quality is characterized in that: The primer set is shown in SEQ ID NOs. 8-9 and SEQ ID NOs. 14-15.

Images