Application of carbohydrate in inhibition of rice tillering growth

Abstract

The invention discloses application of carbohydrates in inhibition of rice tillering growth. Inhibition of rice tillering through the exogenous carbohydrate compound is beneficial to construction of a population structure with a proper size, and high yield of rice is promoted. The invention finds that four carbohydrate compounds have the effect of inhibiting the tillering growth of rice; and the compounds can also be used as food, are non-toxic and harmless to human, and can be safely used as exogenous growth regulating compounds.

Description

Technical Field

[0001] The invention belongs to the field of rice cultivation, and in particular relates to the application of sugar compounds in inhibiting the tillering growth of rice. Background Art

[0002] As the main energy source, carbohydrates can meet the energy metabolism needs of plant growth and development, and can also act as signal molecules similar to hormones. Phenotypic studies of different sugar signals and metabolic mutants have found that carbohydrates not only act as energy in controlling lateral branch growth, but can also regulate the expression patterns of a series of genes and interact with plant hormone signals, playing a regulatory role in the development of plant lateral buds, and their role precedes the hormones in taking a leading role at the bud tip.

[0003] After searching, no reports were found on regulating rice tillering through the action of sugars. Summary of the invention

[0004] In view of the shortcomings of the prior art, the purpose of the present invention is to provide an application of a carbohydrate compound in inhibiting the tillering growth of rice. The present invention found that the minimum effective concentration of trehalose, glucose and sucrose for inhibiting rice tillering is 0.08%, and the minimum effective concentration of glucosamine for inhibiting rice tillering is 0.01%. Spraying exogenous sugar on the leaves will not affect rice tillering, and applying exogenous sugar to the roots is the only way to inhibit rice tillering.

[0005] The technical solution adopted by the present invention to solve the technical problem is:

[0006] In a first aspect, the present invention provides the use of protective carbohydrates in inhibiting tillering growth of rice.

[0007] In a second aspect, the present invention also protects the use of sugar substances in agents for inhibiting the tillering growth of rice.

[0008] In a specific embodiment, the carbohydrate substance is trehalose, glucose, sucrose, glucosamine and the like.

[0009] In a specific embodiment, the effective concentration of trehalose, glucose and sucrose is 0.08% to 0.2%, such as 0.08%, 0.1%, 0.15%, 0.2% and the like.

[0010] In a specific embodiment, the effective concentration of glucosamine is 0.01% to 0.04%, such as 0.01%, 0.02%, 0.04%, etc.

[0011] In a specific embodiment, the agent further contains other active substances that affect the number of rice tillers.

[0012] In a specific embodiment, the reagent further contains other pharmaceutically acceptable carriers or excipients.

[0013] In a third aspect, the present invention protects a method for inhibiting the tillering growth of rice, which comprises applying a sugar substance to the rice.

[0014] In a specific embodiment, the carbohydrate substance is trehalose, glucose, sucrose, glucosamine and the like.

[0015] Beneficial results of the present invention:

[0016] Rice tillering directly affects rice yield. Too many tillers cause the rice population to be closed and poorly ventilated, resulting in reduced rice yield. Inhibiting rice tillering by exogenous sugar compounds is beneficial to building a population structure of appropriate size and promoting high rice yield. The present invention found that four sugar compounds have the effect of inhibiting the growth of rice tillers. In addition, these compounds are also used as food, are non-toxic and harmless to humans, and can be safely used as exogenous growth regulating compounds. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The phenotype comparison of rice treated with different sugars, scale bar = 10 cm. (A) is the phenotype comparison of NIP after 0.02% Tre, Glu and Suc treatment; (B) is the phenotype comparison of NIP after 0.2% Tre, Glu and Suc treatment. Tre: trehalose; Glu: glucose; Suc: sucrose.

[0018] Figure 2 The dynamic changes of rice tillering after different sugar treatments. (A) shows the dynamic changes of NIP tillering after 0.02% Tre, Glu and Suc treatments; (B) shows the dynamic changes of NIP tillering after 0.2% Tre, Glu and Suc treatments. Tre: trehalose; Glu: glucose; Suc: sucrose. P values ​​were calculated by Student's T test, and the significance of the difference was defined as *P<0.05, **P<0.01.

[0019] Figure 3 Comparison of NIP phenotypes of rice treated with different sugars, scale = 10 cm. (A) is the comparison of NIP phenotypes after 0.08% Tre, Glu and Suc treatments, (B) is the tillering dynamics of NIP after 0.08% Tre, Glu and Suc treatments. Tre: trehalose; Glu: glucose; Suc: sucrose. P values ​​were calculated by Student's T test, and significant differences were defined as *P<0.05, **P<0.01.

[0020] Figure 4Phenotypic comparison of rice NIP treated with different concentrations of glucosamine, scale = 10 cm. (A) is the phenotypic comparison of NIP after treatment with 0.01%, 0.02% and 0.04% GlcN, (B) is the tillering dynamics of NIP after treatment with 0.01%, 0.02% and 0.04% GlcN. GlcN: glucosamine. P values ​​were calculated by Student's T test, and significant differences were defined as *P<0.05, **P<0.01.

[0021] Figure 5 Effects of different sugar treatments on the expression of genes related to NIP in rice stem base. Relative expression levels of OsD3 (A), OsD14 (B), OsD53 (C), OsD10 (D), OsCKX4 (E), and OsCKX9 (F) genes at the stem base after 24 h of NIP treatment with 0.08% Glu and 0.01% GlcN. Glu: glucose; GlcN: glucosamine. P values ​​were calculated by Student's T test, and significant differences were defined as *P<0.05, **P<0.01. DETAILED DESCRIPTION

[0022] The technical solution of the present invention is further described below in conjunction with specific implementation schemes. The reagents or instruments used without indicating the manufacturer are regarded as conventional products that can be purchased on the market.

[0023] Example Sugars inhibit rice tillering activity test

[0024] The experiment was set up with different concentrations of different types of exogenous sugar treatments (0.02% glucose, 0.02% trehalose, 0.02% sucrose, 0.2% glucose, 0.2% trehalose, 0.2% sucrose, 0.08% glucose, 0.08% trehalose, 0.08% sucrose, 0.01% glucosamine, 0.02% glucosamine, 0.04% glucosamine) and a control (nutrient solution), and the treatment was uniformly continued for 12 to 20 days.

[0025] The rice cultivation site is the rice cultivation laboratory in the Life Science Building of Nanjing Agricultural University.

[0026] All the seedlings were placed in a light incubator for cultivation, with the light duration set to 15 h, the temperature set to 28° C. / 24° C. (light / darkness), and the relative humidity set to 75%.

[0027] The culture environment is a 96-hole plant hydroponic box (12.7cm×8.7cm×11.4cm) with a nutrient solution volume of 1L, a turnover box (23cm×14cm×9cm) with a nutrient solution volume of 1.5L, and when transplanting, the roots of the rice seedlings are wrapped with a sponge and fixed on a 6-hole foam board (one hole for one seedling, and a spacing of 7cm×6.5cm between rows and plants) and float on the nutrient solution. The nutrient solution is replaced every 4 days, and a corresponding concentration of sugar solution is applied to the roots at the same time when the nutrient solution is replaced.

[0028] The nutrient solution formula is: 1mM KNO3, 1mM NH4Cl, 0.32mM KH2PO4·2H2O, 0.3mM MgCl2·6H2O, 0.67mM CaCl2, 0.3mM Na2SiO3·9H2O, 9μM MnCl2·4H2O, 18.4μM H3BO3, 0.54μMNa2MoO4·2H2O, 0.14μM ZnSO4·7H2O, 0.16μM CuSO4·5H2O, 44.83μM FeSO4·7H2O, 44.76μMNa2EDTA·2H2O.

[0029] After the components of the nutrient solution formula are stirred and mixed, the pH value of the solution is adjusted to 5.5-5.8.

[0030] The treatment method is to continuously observe the growth and development of rice and count the dynamic changes in the number of tillers, select rice plants with normal growth on the 0th day, the 4th day, the 8th day, the 12th day, the 16th day and the 20th day after the exogenous sugar treatment, and count the corresponding number of tillers. After the treatment, the corresponding plant height and root length are measured with a ruler, and the whole plant is photographed and recorded with a single-lens reflex camera to compare the growth differences of rice plants under different sugar treatments.

[0031] The data were analyzed using Excel 2019 for statistical and T-test significance analysis.

[0032] The image processing uses GraphPad Prism8, TBtools, Adobe Illustrator 2020 and Adobe Photoshop2020.

[0033] The difference is that when *P<0.05, it means that the difference has reached a significant level, and when **P<0.01, it means that the difference has reached an extremely significant level.

[0034] Test material: "Nipponbare" rice (NIP)

[0035] The test is described in further detail below with reference to the accompanying drawings.

[0036] 1. Analysis of the regulatory effect of sugars on rice tillering

[0037] In order to clarify the role of different types of exogenous sugars in the growth and development of rice, this experiment, based on the previous preliminary experiments, used low concentrations (0.02%) and high concentrations (0.2%) of trehalose (Tre), glucose (Glu) and sucrose (Suc) to conduct hydroponic experiments on Nipponbare materials.

[0038] like Figure 1 As shown in A, under the treatment of 0.02% concentration of Tre, Glu and Suc, the number of rice tillers after treatment was similar to that of the control. Further analysis of the number of rice tillers during the 0.02% exogenous sugar treatment process found that only the number of tillers of NIP plants treated with 0.02% Tre was significantly lower than that of the control, and the number of tillers of plants treated with other sugars did not reach a significant difference level compared with the control ( Figure 2 A).

[0039] Depend on Figure 1 As can be seen from B, the growth of rice tillers was more strongly inhibited when the exogenous sugar concentration was increased. Figure 2 As shown in B, under the treatment of 0.2% concentration of trehalose, glucose and sucrose, the tiller number of NIP plants was significantly lower than that of the control plants. According to the dynamic changes of rice tiller number, the exogenous sugar that effectively inhibits the tiller number began to significantly inhibit the tiller number 8 days after treatment ( Figure 2 B). The above results show that low concentration (0.02%) of exogenous sugar has no significant effect on rice tillering, and increasing the sugar treatment concentration to 0.2% has a greater effect on rice tillering. Therefore, it can be preliminarily determined that the appropriate exogenous sugar concentration is between 0.02% and 0.2%.

[0040] Based on the results of the previous experiment, this experiment continued to treat NIP with 0.08% concentration of trehalose, glucose and sucrose, and observed and recorded the occurrence of plant tillering. Figure 3 As can be seen from A, 0.08% concentration of trehalose, glucose and sucrose treatment had a significant inhibitory effect on the tillering number of NIP. Figure 3 As shown in B, compared with the control, the tillering of NIP plants was significantly inhibited by 0.08% Tre, Glu, and Suc. 0.08% Glu and 0.08% Suc had a better effect on inhibiting tillering than 0.08% Tre.

[0041] 2. The inhibitory effect of glucosamine on rice tillering

[0042] Glucosamine (GlcN) is an amine sugar in which the hydroxyl group of glucose is replaced by an amino group. It has a wide range of applications in biomedicine, food, health care, industry and agriculture. In order to explore the effect of glucosamine (GlcN) on rice tillering, this study set up a concentration gradient treatment of 0.01%, 0.02%, and 0.04% NIP based on the previous preliminary experiment. Figure 4 As shown in A, compared with the control, different concentrations of GlcN treatment of NIP significantly inhibited plant growth, and the inhibitory effect became more obvious with the increase of treatment concentration. Figure 4 B It can be seen that after NIP was treated with 0.01%, 0.02%, and 0.04% GlcN, the tiller number was significantly reduced compared with the control. The above results show that 0.08% concentration of trehalose, glucose and sucrose and 0.01% glucosamine are involved in the regulation of rice tiller number.

[0043] 3. Changes in gene expression in rice plants after exogenous sugar treatment

[0044] Exogenous sugar treatment has a certain impact on the growth and development of rice plants, and the phenotypic changes of rice are inconsistent with different types and concentrations of exogenous sugar treatment. At present, 0.08% has been identified as the minimum effective concentration of glucose to inhibit rice tillering, and 0.01% is the minimum effective concentration of glucosamine to significantly inhibit tillering. In order to clarify the mechanism of different exogenous sugar treatments on rice tillering inhibition, 0.08% glucose and 0.01% glucosamine were selected for subsequent experiments. NIP plants were treated with corresponding concentrations of exogenous sugar for 24 hours, and the stem bases of the treatment group and the control group were collected for RNA extraction. The extracted RNA was then reverse transcribed to obtain cDNA, which was used as a template for real-time fluorescence quantitative PCR detection experiments. The rice ACTIN gene was used as the internal reference gene in this experiment, and the expression of OsD3, OsD10, OsD14, OsD53, OsCKX4 and OsCKX9 genes at the stem base in NIP was detected.

[0045] like Figure 5 As shown in the figure, compared with the control, the expression level of OsD3 gene detected at the base of rice stems after 0.08% glucose treatment was significantly inhibited by 0.08% glucose. The expression of cytokinin degradation genes OsCKX4 and OsCKX9 was also inhibited by 0.08% glucose, especially the expression of OsCKX9 gene was significantly decreased compared with the control plants, and the expression of OsCKX4 gene had no significant difference after exogenous sugar treatment, only a slight decrease. This indicates that 0.08% glucose may inhibit tillering by affecting the metabolic process of cytokinin and strigolactone. Under 0.01% glucosamine treatment, the expression changes of strigolactone-related genes OsD3, OsD14, OsD10, OsD53 and cytokinin degradation genes OsCKX4 and OsCKX9 at the base of rice stems were significantly different from those after glucose treatment, indicating that the pathway of glucosamine inhibiting rice tillering elongation may be different from that of glucose.

[0046] The protection content of the present invention is not limited to the above embodiments. Without departing from the spirit and scope of the inventive concept, changes and advantages that can be thought of by those skilled in the art are included in the present invention and are protected by the attached claims.

Claims

1. Application of carbohydrates in inhibiting tillering growth of rice.

2. Application of carbohydrates in agents for inhibiting tillering growth of rice.

3. The use according to claim 1 or 2, characterized in that: The sugar substance is trehalose, glucose, sucrose or glucosamine.

4. The use according to claim 3, characterized in that: The effective concentration of the trehalose, glucose and sucrose is 0.08% to 0.2%.

5. The use according to claim 3, characterized in that: The effective concentration of the glucosamine is 0.01% to 0.04%.

6. The use according to claim 2, characterized in that: The reagent also contains other active substances that affect the tillering quantity of rice.

7. The use according to claim 2, characterized in that: The reagent may also contain other pharmaceutically acceptable carriers or excipients.

8. A method for inhibiting tillering growth of rice, the method comprising applying sugar substances to the rice.

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