A method for spraying exogenous brassinolide for millet

Abstract

The present application discloses a method for spraying exogenous brassinolide for millet, comprising the following steps: step S1) formula preparation, preparing a 24-epibrassinolide spraying solution with a concentration of 0.10-0.25 mg / L, mixing brassinolide raw powder with a biological additive in a mass ratio of 1:2-1:4, wherein the biological additive is a complex of chitosan and a water-soluble polysaccharide; step S2) solution preparation, diluting the prepared mixture with a buffer solution with a pH value of 6.5-7.2, adding 0.02-0.05% of a surfactant, and stirring evenly. This application innovatively established a differentiated formulation system based on the plant characteristics of millet. By screening specific biological auxiliary agent complexes, the bioavailability of brassinolide was significantly improved. For upright millet, a composite carrier of chitosan and carrageenan was used, combined with the synergistic effect of pyrrolidone formic acid, which greatly improved the operation efficiency of the drug solution in the vascular bundle of the stem, effectively achieving consistent control of plant height and synchronous regulation of ear development.

Description

Technical Field

[0001] The invention relates to the technical field of fertilization, in particular to a method for spraying exogenous brassinolide for millet. Background Art

[0002] As an important type of plant growth regulator, brassinolide (BL) has many physiological functions in the growth and development of millet, such as promoting cell elongation, inducing vascular tissue differentiation, improving stress resistance, and regulating source-sink relationship. At present, the application of exogenous brassinolide in millet production mainly includes seed soaking, root infiltration and foliar spraying. Among them, foliar spraying is widely used in production practice due to its advantages such as convenient operation, rapid absorption and significant regulation effect. In the existing technology, a single concentration (0.1-0.2 mg / L) of 24-epibrassinolide solution is usually used for spraying at a specific growth period, and a simple surfactant is used in combination to improve the wettability and permeability of the solution on the leaf surface, which promotes the growth and development of millet and the formation of yield to a certain extent.

[0003] However, the existing brassinolide spraying technology has many technical bottlenecks and application limitations: first, it fails to fully consider the differences in morphological structure, physiological characteristics and response mechanism of millet of different plant types, and the use of a unified formula leads to insufficient efficacy and even negative effects; second, there is a lack of precise application schemes based on the growth process of millet, especially in key growth periods such as tillering, jointing, ear differentiation and heading, and a systematic concentration progression system and spraying timing judgment criteria have not been established; third, the selection and ratio of biological adjuvants lack theoretical guidance, and the carrier function and synergistic effect of biomaterials such as chitosan and water-soluble polysaccharides have not been fully utilized; in addition, the existing technology is not precise enough in regulating the physical and chemical properties of the liquid such as pH value and ionic strength, resulting in poor stability of the active ingredients and low utilization rate; finally, there is a lack of systematic response strategies for special weather conditions such as rainy, high temperature, drought and strong wind, which seriously affects the actual application effect and promotion value of brassinolide. For example, the existing patent with publication number CN115968730A discloses a method for spraying 2,4-epibrassinolide on wheat, which sprays epibrassinolide twice at intervals during the rooting stage + grain filling stage or the jointing stage + grain filling stage, with a spraying concentration of 0.075 mg / L, 30 L of solution per mu, and 0.1% Tween 20 is added as a surface lubricant during spraying, which is conducive to high-yield and high-efficiency production of wide-area sowing wheat and reduces the impact on the environment. The application (CN115968730A) has the technical defects of the above-mentioned brassinolide spraying technology.

[0004] Therefore, it is urgent to develop a precise spraying technology system of exogenous brassinolide based on the physiological characteristics of millet with different plant types, adapting to the needs of different growth periods, and having the ability to defend against special weather. This technology system should include optimized formula components, perfect spraying technology, precise dosage control and supporting cultivation measures to comprehensively improve the application effect and scientific and technological contribution rate of brassinolide in millet production. Summary of the invention

[0005] The purpose of the present invention is to provide a method for spraying exogenous brassinolide on millet, so as to solve the defects of the brassinolide spraying technology in the prior art and comprehensively improve the application effect of brassinolide in millet production.

[0006] To achieve the above object, the present invention provides the following technical solution, a method for spraying exogenous brassinolide on millet, comprising the following steps:

[0007] Step S1) Recipe preparation:

[0008] A 24-epi-brassinolide spray solution with a concentration of 0.10-0.25 mg / L is prepared, and the brassinolide raw powder and the biological additive are mixed in a mass ratio of 1:2-1:4, wherein the biological additive is a complex of chitosan and a water-soluble polysaccharide;

[0009] Step S2) Solution preparation:

[0010] The prepared mixture was diluted with a buffer solution having a pH value of 6.5-7.2, 0.02-0.05% of a surfactant was added, and stirred evenly;

[0011] Step S3) Spraying during the growth period:

[0012] S31) Tillering stage:

[0013] During the peak tillering period, spray at a temperature of 0-25°C with a spraying concentration of 0.10-0.15 mg / L;

[0014] S32) Jointing stage:

[0015] Spray when the first internode is elongated, with a spray concentration of 0.12-0.18 mg / L;

[0016] S33) Ear differentiation stage:

[0017] Spray at the beginning of spikelet differentiation, with a spraying concentration of 0.14-0.22 mg / L;

[0018] S34) Heading period:

[0019] Spray at the heading stage with a concentration of 0.15-0.25 mg / L;

[0020] Among them, the spraying conditions are selected from 2 hours before sunrise to sunrise or from sunset to 1 hour after sunset, the ambient temperature is controlled at 16-27℃, the relative humidity is controlled at 60-85%, the wind speed is less than 2m / s, and no rainfall is expected within 24 hours.

[0021] Preferably, in step S1, the formula preparation is selected as follows according to the millet plant type:

[0022] When the millet is an upright plant type, the concentration of brassinolide is 0.15-0.20 mg / L, chitosan and carrageenan are selected with a mass ratio of 3:1, and 0.03% pyrrolidone formic acid is added;

[0023] When it is tillering millet, the concentration of brassinolide is 0.10-0.15 mg / L, chitosan and gum arabic are selected with a mass ratio of 2:1, and 0.02% betaine is added;

[0024] When it is a hybrid millet variety, the brassinolide concentration is 0.20-0.25 mg / L, chitosan and xanthan gum are selected with a mass ratio of 4:1, and 0.04% amino oligosaccharide is added.

[0025] Preferably, in step S1, the method for preparing the biological additive comprises:

[0026] Step S11) Preparation of chitosan solution:

[0027] Chitosan with a deacetylation degree of 85-95% and a molecular weight of 50-150 kDa was selected and dissolved in 1% acetic acid solution;

[0028] Step S12) Preparation of water-soluble polysaccharide solution:

[0029] Dissolve the water-soluble polysaccharide in deionized water to prepare a 2-5% aqueous solution;

[0030] Step S13) Composite preparation:

[0031] At 25-30° C., the solution obtained in step S11 and the solution obtained in step S12 are mixed according to a set ratio.

[0032] Preferably, in step S2, the buffer is selected from:

[0033] When pH < 7.0, use a mixture of citric acid and disodium hydrogen phosphate;

[0034] When pH ≥ 7.0, use a mixture of potassium dihydrogen phosphate and sodium hydroxide;

[0035] The ionic strength of the buffer solution is 0.1-0.2 mol / L, and when the pH changes by ±0.5, the buffer capacity is not less than 0.02 mol / L.

[0036] Preferably, in step S2, the surfactant must satisfy:

[0037] There is no precipitation, turbidity or stratification within 48 hours of mixing with brassinolide. The surface tension of the aqueous solution can be reduced to 25-35mN / m, the critical micelle concentration is 0.015-0.025%, and the retention rate of the active ingredient is greater than 85% after 2mm precipitation.

[0038] Preferably, in step S31, the spraying during the tillering stage includes:

[0039] The first spraying is done when the base of the tillering node begins to swell, and the second spraying is done 10-15 days later;

[0040] Humidify the stem base 2 hours before the first spraying;

[0041] The spraying height is 30-40cm from the leaf surface, and the spraying pressure is 0.15-0.20MPa;

[0042] Add 0.02-0.03% water-soluble boron fertilizer and 0.01-0.02% cytokinin to the spray solution;

[0043] Avoid watering within 4-6 hours after spraying and keep the relative moisture content of the soil at 65-70%.

[0044] Preferably, in step S32, the jointing stage spraying process includes:

[0045] The first spraying is carried out when the length of the first internode reaches 0.5-1cm;

[0046] 5-7 days after the first spraying, measure the mechanical strength of the stems, and spray a second time when the mechanical strength of the stems is less than 85% of the standard value;

[0047] When spraying, add 0.02-0.04% potassium fertilizer and 0.01-0.02% salicylic acid to promote stem thickening and lignin formation;

[0048] Ensure that the coverage of the spray on the stem surface reaches more than 90%, the coefficient of variation of droplet distribution uniformity is less than 10%, and the unit area dosage is controlled at 45-60L / hm²;

[0049] No mechanical operation should be carried out within 12 hours after spraying, and the soil moisture should be maintained at 75-80% of the field water holding capacity.

[0050] Preferably, in step S33, the spraying process during the ear differentiation period includes:

[0051] Select the time when the spikelet length is 1-2 mm for the first spraying;

[0052] 3-5 days after the first spraying, the leaf chlorophyll fluorescence parameter Fv / Fm value is used to determine whether a second spraying is needed. When the Fv / Fm value is lower than 0.75, the second spraying is performed.

[0053] The dosage of liquid per unit area is 45-55L / hm², and 0.02-0.03% putrescine and 0.01-0.02% gibberellin are added at the same time for synergistic promotion;

[0054] During the spraying process, samples were taken every 50m² to check the ear distribution to ensure that the liquid was evenly distributed;

[0055] When spraying on sunny days, use a shade net with a shading rate of 30-40% within 2 hours after spraying, and maintain the relative humidity in the field at 70-80%.

[0056] Preferably, in step S34, the heading period spraying process includes:

[0057] The first spraying should be done when the flag leaves are fully expanded but the ear has not yet emerged;

[0058] 4-6 days after the first spraying, observe the ear length and spray a second time when the ear length reaches 2-3 cm;

[0059] Add 0.03-0.05% of putrescine and 0.02-0.03% of calcium and magnesium chelating agent to the spray solution;

[0060] Control the spray particle size to 60-90μm, and the amount of liquid used per unit area to 50-60L / hm²;

[0061] When spraying on cloudy days, ensure that the coverage of the liquid on the plant surface reaches more than 85%, the coefficient of variation of the uniformity of the liquid deposition on the ear is less than 15%, and the relative humidity of the canopy is maintained at 75-85% after spraying;

[0062] Monitor the leaf moisture content within 24 hours after spraying, and add additional leaf humidification when it is below 85%.

[0063] Preferably, the spraying adjustment measures under special weather conditions include:

[0064] Rainy weather treatment: When it rains continuously, increase the concentration of the liquid medicine by 15-20%, add 0.02-0.03% of vine extension agent and 0.01-0.02% of pyrrolidone compounds, choose the interval between rains for spraying, and reduce the dosage to 80-85% of the standard dosage;

[0065] High temperature weather treatment: When the temperature is higher than 32℃, spray before the morning dew dries up, reduce the concentration of brassinolide to 70-80% of the standard concentration, add 0.02-0.03% betaine as a protective agent, and use a shade net to shade after spraying, with the shading rate controlled at 40-50%;

[0066] Drought stress treatment: Supplementary spraying is carried out from sunset to after sunset, adding 0.02-0.03% moisturizer and 0.01-0.02% potassium humate. After spraying, cover the ground with straw or biodegradable film in time, increase the number of spraying, and reduce the amount used each time to 60-70% of the standard amount;

[0067] Treatment for windy weather: When the wind speed exceeds 3m / s, suspend spraying. When the wind speed drops below 3m / s, choose to spray row by row in the leeward direction. Re-spray the sprayed area once within 12 hours after the wind speed decreases. During the supplementary spraying, increase the brassinolide concentration by 10-15%.

[0068] Compared with the prior art, the present invention has the following beneficial effects:

[0069] 1) This application innovatively established a differentiated formulation system based on millet plant characteristics. By screening specific biological adjuvant complexes, the bioavailability of brassinolide was significantly improved. For erect millet, a composite carrier of chitosan and carrageenan was used, combined with the synergistic effect of pyrrolidone formic acid, which greatly improved the operation efficiency of the liquid medicine in the vascular bundle of the stem. For tillering millet, a composite system of chitosan and gum arabic was selected, combined with the osmotic regulation of betaine, which significantly promoted the accumulation and transport of hormones in the tillering node and significantly improved the effective tillering ear formation rate. For hybrid varieties, a carrier system of chitosan and xanthan gum was used, combined with the signal induction effect of amino oligosaccharides, to effectively achieve consistent control of plant height and synchronous regulation of ear development;

[0070] 2) This application has established a complete growth period precision spraying technology system. By analyzing the sensitivity and response characteristics of millet to brassinolide at different growth periods, a scientific concentration progression scheme has been designed. By adding water-soluble boron fertilizer and cytokinin at the tillering stage, the tillering ear rate is significantly improved. The synergistic effect of potassium fertilizer and salicylic acid at the jointing stage greatly improves the mechanical strength of the stem and significantly reduces the lodging rate. The compound use of putrescine and gibberellin at the ear differentiation stage significantly improves the uniformity of ear differentiation. The regulation of calcium and magnesium chelators at the heading stage effectively promotes the grain filling rate and improves the quality of millet.

[0071] 3) This application has established a response mechanism for special weather conditions, and has developed systematic control measures for different types of adverse weather. Under rainy weather conditions, by optimizing the concentration of the liquid medicine, adding vine extension agents and pyrrolidone compounds, the uniform distribution and persistence of the liquid medicine on the leaf surface are significantly improved. Under high temperature conditions, the strategy of spraying before the morning dew has dried, combined with the protective effect of betaine, effectively slowed down the downward trend of the photosynthetic rate of the plants. Under drought stress, spraying in the evening and adding moisturizers and potassium humate, combined with surface covering measures, significantly enhanced the drought resistance of the plants. Under windy conditions, the uniformity of the application and the persistence of the efficacy of the medicine are effectively guaranteed by scientifically adjusting the direction and timing of spraying;

[0072] 4) In terms of spraying technology, this application has established a complete quality control system by accurately controlling technical indicators such as spray coverage, droplet distribution uniformity and deposition rate. At the same time, it innovatively uses indicators such as chlorophyll fluorescence parameters and stem mechanical strength as the basis for judging secondary spraying, and through fixed-point sampling and analysis, it significantly improves the scientificity and controllability of the spraying effect. Studies have shown that this technology has higher efficacy and better economy than traditional spraying methods;

[0073] 5) The biological additive compound system developed in this application has a significant synergistic effect. Through the compounding of chitosan and different water-soluble polysaccharides, not only the stability and bioavailability of brassinolide are improved, but also the stress resistance of the plant is enhanced. Among them, through the optimization design of the deacetylation degree and molecular weight of chitosan, combined with strict pH value and ionic strength control, the liquid medicine system has excellent physical and chemical stability and biocompatibility. By adding synergists with different functions, such as salicylic acid, putrescine substances, calcium and magnesium chelators, etc., all-round regulation of millet growth and development is achieved, achieving the goal of improving quality and increasing production. Field trials have shown that the use of this technical system can significantly improve millet yield and quality, and has good application prospects and promotion value. BRIEF DESCRIPTION OF THE DRAWINGS

[0074] Figure 1 This is a data summary table of the effects of different concentrations of brassinolide treatment on millet growth and physiological indicators in Experimental Example 1 of the present invention;

[0075] Figure 2 This is a data summary table of the differences in responses of millet of different plant types to brassinolide concentrations in Experimental Example 1 of the present invention;

[0076] Figure 3 This is a data summary table showing the effects of different ratios on the physicochemical properties of the brassinolide preparation in Example 1 of the present invention;

[0077] Figure 4A data summary table showing the effects of spraying treatments at different growth stages on millet growth and yield in Example 2 of the present invention;

[0078] Figure 5 This is a data summary table showing the effects of different formulations in Example 3 of the present invention on the growth and physiological indicators of upright plant type millet;

[0079] Figure 6 This is a data summary table of the effects of different formulations in Example 3 of the present invention on the tillering characteristics of tillering millet;

[0080] Figure 7 A data summary table showing the effects of different formulations in Example 3 of the present invention on the uniformity of growth of hybrid varieties;

[0081] Figure 8 This is a data summary table of the effects of different surfactants on the performance of the brassinolide preparation in Example 4 of the present invention;

[0082] Fig. 9 This is a data summary table showing the effects of different preparation processes on the performance of the bio-adjuvant composite in Example 4 of the present invention;

[0083] Fig.10 is a flow chart of the spraying method of the present invention;

[0084] Fig.11 The present invention is a flow chart of the method for preparing the biological additive. DETAILED DESCRIPTION

[0085] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0086] In the description of the invention, it should be noted that the terms "upper", "lower", "inner", "outer", "front end", "rear end", "two ends", "one end", "the other end" and the like indicate positions or positional relationships based on the positions or positional relationships shown in the drawings, and are only for the convenience of describing the invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operate in a specific orientation, and therefore cannot be understood as limiting the invention. In addition, the terms "first" and "second" are used for descriptive purposes only and cannot be understood as indicating or implying relative importance.

[0087] In the description of the invention, it should be noted that, unless otherwise clearly specified and limited, the terms "installed", "provided with", "connected", etc. should be understood in a broad sense. For example, "connected" can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection or an electrical connection; it can be a direct connection, or it can be indirectly connected through an intermediate medium, or it can be the internal communication of two components. For ordinary technicians in this field, the specific meanings of the above terms in the present invention can be understood according to specific circumstances.

[0088] In the description of the invention, it should be noted that the execution order of the steps is not limited by the serial numbers, and the order of some steps can be changed, the steps can be executed simultaneously, the steps can be executed separately, etc., all of which are within the protection scope of this application.

[0089] See also Figure 1-11 The present invention provides a method for spraying exogenous brassinolide on millet, comprising the following steps:

[0090] Step S1) Recipe preparation:

[0091] A 24-epi-brassinolide spray solution with a concentration of 0.10-0.25 mg / L is prepared, and the brassinolide raw powder and the biological additive are mixed in a mass ratio of 1:2-1:4, wherein the biological additive is a complex of chitosan and a water-soluble polysaccharide;

[0092] Step S2) Solution preparation:

[0093] The prepared mixture was diluted with a buffer solution having a pH value of 6.5-7.2, 0.02-0.05% of a surfactant was added, and stirred evenly;

[0094] Step S3) Spraying during the growth period:

[0095] S31) Tillering stage:

[0096] During the peak tillering period, spray at a temperature of 20-25°C with a spraying concentration of 0.10-0.15 mg / L;

[0097] S32) Jointing stage:

[0098] Spray when the first internode is elongated, with a spray concentration of 0.12-0.18 mg / L;

[0099] S33) Ear differentiation stage:

[0100] Spray at the beginning of spikelet differentiation, with a spraying concentration of 0.14-0.22 mg / L;

[0101] S34) Heading period:

[0102] Spray at the heading stage with a concentration of 0.15-0.25 mg / L;

[0103] Among them, the spraying conditions are selected from 2 hours before sunrise to sunrise or from sunset to 1 hour after sunset, the ambient temperature is controlled at 16-27℃, the relative humidity is controlled at 60-85%, the wind speed is less than 2m / s, and no rainfall is expected within 24 hours.

[0104] Specifically, by setting a reasonable concentration range (0.10-0.25 mg / L), biological additive ratio (1:2-1:4) and spraying strategy at different growth stages, the application of brassinolide was systematized and standardized.

[0105] As an additional explanation, 24-epibrassinolide is a steroidal compound with a specific spatial configuration and is the most active brassinolide discovered so far. The configuration of the carbon atom at position 24 in its molecular structure plays a key role in its activity; in this application, chitosan and water-soluble polysaccharides are complex bio-auxiliaries, which are auxiliary materials used to improve the bioavailability of pesticides or growth regulators; chitosan has good biocompatibility and film-forming properties and can be used as a drug carrier; water-soluble polysaccharides are hydrophilic macromolecular polysaccharide compounds, which are used in this application to improve the physical and chemical properties of the drug solution.

[0106] As an additional note, cells divide vigorously during the tillering stage and are sensitive to exogenous hormones; the jointing stage is the critical period for the transition from vegetative growth to reproductive growth, and the ear differentiation stage is the critical stage for the formation and differentiation of young ear primordia, which is particularly sensitive to changes in hormone levels; the heading stage is the decisive stage for grain formation, and hormone levels directly affect the filling efficiency.

[0107] Specifically, the spraying conditions are selected from 2 hours before sunrise to sunrise or from sunset to 1 hour after sunset. For example, the spraying time is selected before 8 am or after 6 pm. The temperature is relatively low and the humidity is high during these two periods, which is conducive to the absorption of the liquid medicine. In different seasons, in spring, you can choose 1-2 hours before sunrise or 0.5-1 hour after sunset, in summer, you can choose 1.5-2.5 hours before sunrise or 1-1.5 hours after sunset, and in autumn, you can choose 1-1.5 hours before sunrise or 0.5-1 hour after sunset.

[0108] Specifically, spraying conditions should be adjusted appropriately under special weather conditions. The morning spraying time can be appropriately extended on cloudy days, the morning spraying time should be advanced in hot weather, spraying in the evening is preferred in drought conditions, and spraying should be carried out after the dew has decreased in foggy weather.

[0109] In some embodiments, in step S1, the formula preparation is selected according to the millet plant type as follows:

[0110] When the millet is an upright plant type, the concentration of brassinolide is 0.15-0.20 mg / L, chitosan and carrageenan are selected with a mass ratio of 3:1, and 0.03% pyrrolidone formic acid is added;

[0111] When it is tillering millet, the concentration of brassinolide is 0.10-0.15 mg / L, chitosan and gum arabic are selected with a mass ratio of 2:1, and 0.02% betaine is added;

[0112] When it is a hybrid millet variety, the brassinolide concentration is 0.20-0.25 mg / L, chitosan and xanthan gum are selected with a mass ratio of 4:1, and 0.04% amino oligosaccharide is added.

[0113] Experimental Example 1: Optimization experiment of the spraying concentration of brassinolide

[0114] 1. Test materials:

[0115] Millet varieties Jingu 21 (representative upright plant type variety), Jingu 59 (representative tillering type variety) and Changzagu 466 (representative hybrid variety) were selected as test materials. 24-Epibrassinolide (purity ≥ 98%) was prepared.

[0116] 2. Test location and time:

[0117] The experiments were conducted in three representative ecological zones, namely, the Lishi District Experiment Station of Luliang City (37°08'N, 111°48'E, 1100m above sea level, Loess Plateau), the Gaoping City Experiment Station of Jincheng City (35°47'N, 112°55'E, 800m above sea level, Taihang Mountain Area), and the Yongji City Experiment Station of Yuncheng City (34°52'N, 110°27'E, 380m above sea level, Fenwei Plain Area), and the experiments will be carried out for three consecutive years (2021-2023).

[0118] 3. Experimental design:

[0119] (1) Potted plant test:

[0120] Use a plastic pot with a diameter of 25 cm and a height of 30 cm and fill it with 10 kg of soil;

[0121] Sow 10 seeds per pot and thin out to 6 plants after emergence;

[0122] Set 6 concentration gradients: 0.05, 0.10, 0.15, 0.20, 0.25, 0.30 mg / L;

[0123] Each treatment was set up with 4 replicates, arranged in random blocks;

[0124] (2) Field trials:

[0125] The plot area is 20m²;

[0126] The row spacing is 30cm and the plant spacing is 10cm;

[0127] The concentration gradient was the same as the pot experiment;

[0128] A randomized block design was used with 3 replications.

[0129] 4. Cultivation management:

[0130] Field management was carried out according to the local high-yield cultivation model, and fertilizer and water management remained consistent, with spraying treatments carried out during the tillering stage, jointing stage, ear differentiation stage, and heading stage respectively.

[0131] 5. Measurement indicators:

[0132] (1) Morphological indicators:

[0133] Plant height: measured with a ruler, 20 plants were measured in each plot;

[0134] Stem diameter: measured with a vernier caliper, 20 plants were measured in each plot;

[0135] Tillering number: 20 plants were surveyed in each plot;

[0136] Leaf area: measured using CI-203 laser leaf area meter;

[0137] (2) Physiological indicators:

[0138] Photosynthetic parameters: Li-6400XT photosynthetic instrument was used to measure net photosynthetic rate, stomatal conductance and intercellular CO 2 concentration;

[0139] Chlorophyll content: measured using a SPAD-502 chlorophyll meter;

[0140] Endogenous hormone levels: Liquid chromatography-mass spectrometry was used to determine the contents of IAA, GA3, ZR and ABA.

[0141] 6. Test results:

[0142] (1) Impact on plant growth:

[0143] 0.05mg / L treatment: compared with the control, the growth indicators did not increase significantly (P>0.05);

[0144] 0.10-0.25 mg / L treatment: plant height, stem diameter, tiller number and leaf area were significantly higher than those of the control (P<0.05);

[0145] 0.30mg / L treatment: Growth inhibition occurred, and plant height decreased by 8-12% compared with 0.25mg / L treatment;

[0146] (2) Impact on photosynthetic characteristics:

[0147] In the range of 0.10-0.25 mg / L, the net photosynthetic rate increased with the concentration;

[0148] The 0.20 mg / L treatment reached the maximum value, which was 25-30% higher than the control;

[0149] The photosynthetic parameters showed a downward trend in the 0.30 mg / L treatment;

[0150] (3) Changes in endogenous hormone levels:

[0151] The 0.15-0.20 mg / L treatment significantly increased the levels of endogenous IAA and GA3;

[0152] Treatments above 0.25 mg / L resulted in a significant increase in ABA content;

[0153] Optimal treatment can increase the ZR content by 40-50%.

[0154] 7. Conclusion:

[0155] Therefore, the results of three years of multi-point tests show that the optimal spraying concentration range of brassinolide is 0.10-0.25 mg / L.

[0156] The optimum concentration for upright plant varieties is 0.15-0.20 mg / L;

[0157] The optimum concentration for tillering varieties is 0.10-0.15 mg / L;

[0158] The optimum concentration for hybrid varieties is 0.20-0.25 mg / L;

[0159] Within this concentration range, it can significantly promote millet growth and development, improve photosynthetic efficiency, and optimize endogenous hormone levels. When it is lower than 0.10 mg / L, the growth-promoting effect is not significant; when it is higher than 0.25 mg / L, an inhibitory effect may occur. This result shows good consistency and stability in experiments in different ecological zones and different years.

[0160] In Experimental Example 1, the results of the physical and chemical index determination of the optimization test of the brassinolide spraying concentration are as follows:

[0161] Table 1 Effects of different concentrations of brassinolide on millet growth and physiological indicators (data are the average values ​​of three-year three-point experiments)

[0162]

[0163] Notes to Table 1:

[0164] 1. The data are the average values ​​of three varieties (Jingu 21, Jingu 59 and Changzagu 466);

[0165] 2.* indicates significant difference compared with the control group (P<0.05), ** indicates extremely significant difference compared with the control group (P<0.01);

[0166] 3.FW means fresh weight.

[0167] Table 2 Differences in response of millet of different plant types to brassinolide concentrations (data measured at heading stage)

[0168]

[0169] Notes to Table 2:

[0170] 1. The increase data are percentages relative to the control;

[0171] 2. The total amount of endogenous hormones includes the comprehensive measured values ​​of IAA, GA3 and ZR;

[0172] 3. Data are the average of three-year three-point tests.

[0173] Example 1: Biological additive ratio optimization test

[0174] 1. Test materials:

[0175] (1) Brassinolide: 24-epibrassinolide (purity ≥ 98%);

[0176] (2) Chitosan: deacetylation degree 90%, molecular weight 100 kDa;

[0177] (3) Water-soluble polysaccharides: carrageenan, gum arabic, xanthan gum (all analytically pure);

[0178] 2. Experimental design:

[0179] Set up 5 ratio treatments:

[0180] T1: Brassinolide: biological additive = 1:1

[0181] T2: Brassinolide: biological additive = 1:2

[0182] T3: Oleocin lactone: biological additive = 1:3

[0183] T4: Brassinolide: biological additive = 1:4

[0184] T5: Brassinolide: biological additive = 1:5

[0185] Each treatment was repeated 4 times.

[0186] 3. Measurement indicators:

[0187] (1) Physical and chemical properties:

[0188] Mixing uniformity, dispersion stability, particle size distribution, drug loading, and drug release rate;

[0189] (2) Biological effect indicators:

[0190] Leaf absorption rate, tissue distribution, and duration of efficacy;

[0191] 5. Test results:

[0192] See the table below:

[0193] Table 3 Effects of different ratios on the physicochemical properties of brassinolide preparations

[0194]

[0195] Example 2: Verification test of spraying effect in different stages during growth period

[0196] 1. Purpose of the test:

[0197] Verify the rationality of spraying plans at different growth stages and determine the optimal spraying period and concentration.

[0198] 2. Test materials:

[0199] (1) Test variety: Jingu 21 (upright plant type);

[0200] (2) Test agent: Brassinolide preparation optimized according to Example 1.

[0201] 3. Experimental treatment:

[0202] T1: spraying only during the tillering stage;

[0203] T2: spraying only at the jointing stage;

[0204] T3: spraying only during the ear differentiation period;

[0205] T4: spraying only at heading stage;

[0206] T5: spraying during the entire growth period (tillering stage, jointing stage, ear differentiation stage, heading stage);

[0207] CK: clean water spray;

[0208] 4. Pesticide concentration at each period:

[0209] Tillering stage: 0.10-0.15 mg / L;

[0210] Jointing stage: 0.12-0.18 mg / L;

[0211] Ear differentiation stage: 0.14-0.22 mg / L;

[0212] Heading stage: 0.15-0.25 mg / L;

[0213] 5. Test results:

[0214] See the table below:

[0215] Table 4 Effects of spraying treatments at different growth stages on millet growth and yield

[0216]

[0217] Notes to Table 4:

[0218] * indicates significant difference compared with the control group (P<0.05), ** indicates extremely significant difference compared with the control group (P<0.01).

[0219] From Table 3 and Table 4, we can see that:

[0220] (1) Example 1 shows that the optimal ratio of brassinolide to biological additive is 1:2-1:4. Within this range, the preparation has good physical and chemical properties and biological effects. When the ratio is lower than 1:2, the stability of the preparation is poor; when it is higher than 1:4, the release rate is too slow.

[0221] (2) Example 2 proves that the spraying scheme in different growth stages is significantly better than spraying in a single period, and the full growth period treatment (T5) increases yield by 20.7% compared with the control. The progressive concentration design is adopted in each period, which conforms to the law of crop growth and development and achieves the purpose of improving quality and increasing yield.

[0222] Specifically, a personalized formula system was established for millets of different plant types. The upright millet uses a chitosan and carrageenan (3:1) ratio, combined with pyrrolidone formic acid, which fully utilizes the film-forming property of carrageenan and the property of pyrrolidone formic acid in promoting vascular bundle development; the tillering millet uses a chitosan and gum arabic (2:1) ratio, combined with the osmotic regulation of betaine, which significantly improves the hormone response of the tillering node; the hybrid variety uses a chitosan and xanthan gum (4:1) ratio, and achieves synchronous growth regulation through the signal induction of amino oligosaccharides. This differentiated formula design fully considers the physiological characteristics and growth needs of different plant types, greatly improving the accuracy and effectiveness of regulation.

[0223] Example 3: Experiment on optimizing formula of brassinolide in millet of different plant types

[0224] 1. Test materials:

[0225] (1) Test varieties:

[0226] Upright plant type: Jingu 21, Jingu 16;

[0227] Tillering type: Jingu 59, Changnong 47;

[0228] Hybrid varieties: Changzagu 466, Zhangzagu 5;

[0229] (2) Experimental drugs:

[0230] 24-Epibrassinolide (purity ≥98%);

[0231] Chitosan (90% deacetylation);

[0232] Carrageenan, gum arabic, xanthan gum;

[0233] Pyrrolidone carboxylic acid, betaine, amino oligosaccharides;

[0234] 2. Experimental design:

[0235] Each plant type was set up as follows:

[0236] (1) Upright plant type treatment group:

[0237] D1: brassinolide 0.15 mg / L + chitosan and carrageenan (2:1) + 0.03% pyrrolidone formic acid;

[0238] D2: brassinolide 0.18 mg / L + chitosan and carrageenan (3:1) + 0.03% pyrrolidone formic acid;

[0239] D3: brassinolide 0.20 mg / L + chitosan and carrageenan (4:1) + 0.03% pyrrolidone formic acid;

[0240] D4: single brassinolide 0.18 mg / L (control);

[0241] (2) Tillering type treatment group:

[0242] F1: brassinolide 0.10 mg / L + chitosan and gum arabic (1:1) + 0.02% betaine;

[0243] F2: brassinolide 0.12 mg / L + chitosan and gum arabic (2:1) + 0.02% betaine;

[0244] F3: brassinolide 0.15 mg / L + chitosan and gum arabic (3:1) + 0.02% betaine;

[0245] F4: single brassinolide 0.12 mg / L (control);

[0246] (3) Hybrid treatment group:

[0247] H1: brassinolide 0.20 mg / L + chitosan and xanthan gum (3:1) + 0.04% amino oligosaccharide;

[0248] H2: brassinolide 0.22 mg / L + chitosan and xanthan gum (4:1) + 0.04% amino oligosaccharide;

[0249] H3: brassinolide 0.25 mg / L + chitosan and xanthan gum (5:1) + 0.04% amino oligosaccharide;

[0250] H4: single brassinolide 0.22 mg / L (control);

[0251] 3. Measurement indicators:

[0252] (1) Plant type characteristic indicators:

[0253] The number of vascular bundles in the stem, the number of tillers and ear formation rate, the consistency of plant height, and ear characteristics;

[0254] (2) Physiological indicators:

[0255] Stem mechanical strength, changes in hormone content, photosynthetic parameters, and root activity;

[0256] 4. Test results:

[0257] See the table below:

[0258] Table 5 Effects of different formulas on growth and physiological parameters of upright millet

[0259]

[0260] Table 6 Effects of different formulations on tillering characteristics of tillering millet

[0261]

[0262] Table 7 Effects of different formulas on growth uniformity of hybrid varieties

[0263]

[0264] Notes to Table 6-7:

[0265] 1. FW means fresh weight;

[0266] 2. CV is the coefficient of variation. The smaller the value, the higher the uniformity.

[0267] 3. The data are the average of the two tested varieties;

[0268] 4. The yield index was calculated with the control treatment as 1.00;

[0269] 5. The value after ± indicates the standard deviation;

[0270] Therefore, according to Table 5, Table 6, and Table 7, we can know that:

[0271] (1) Upright millet:

[0272] The optimal formula was D2 treatment, which was 0.18 mg / L brassinolide + chitosan and carrageenan (3:1) + 0.03% pyrrolidone formic acid. This formula significantly enhanced the mechanical strength of the stems, improved the level of vascular bundle development, and improved lodging resistance.

[0273] (2) Tillering millet:

[0274] The best formula was the F2 treatment, which was 0.12 mg / L brassinolide + chitosan and gum arabic (2:1) + 0.02% betaine. This formula significantly increased the tillering rate, enhanced root activity, and promoted the accumulation of hormones at the tillering node.

[0275] (3) Hybrid varieties:

[0276] The optimal formula was H2 treatment, i.e. 0.22 mg / L brassinolide + chitosan and xanthan gum (4:1) + 0.04% amino oligosaccharide. This formula significantly improved the uniformity of the population, increased the fruiting rate, and achieved synchronous growth.

[0277] As an additional note, carrageenan has a unique spiral structure, which is conducive to the formation of a uniform film layer of the drug solution on the surface of the upright stems; the addition of pyrrolidone formic acid can promote cell wall relaxation and enhance vascular bundle development; gum arabic has good emulsifying properties, which is conducive to the uniform distribution of the drug solution in the tillering nodes; the addition of betaine can regulate osmotic pressure and promote hormone transport between nodes; xanthan gum has a special network structure and achieves a sustained release effect; the addition of amino oligosaccharides can induce plant defense responses and enhance resistance.

[0278] In some embodiments, in step S1, the method for preparing a biological additive comprises:

[0279] Step S11) Preparation of chitosan solution:

[0280] Chitosan with a deacetylation degree of 85-95% and a molecular weight of 50-150 kDa was selected and dissolved in 1% acetic acid solution;

[0281] Step S12) Preparation of water-soluble polysaccharide solution:

[0282] Dissolve the water-soluble polysaccharide in deionized water to prepare a 2-5% aqueous solution;

[0283] Step S13) Composite preparation:

[0284] At 25-30° C., the solution obtained in step S11 and the solution obtained in step S12 are mixed according to a set ratio.

[0285] As an additional explanation, acetic acid can provide a suitable pH environment for protonation and dissolution of chitosan; deionized water should use pure water with a conductivity of less than 0.1 μS / cm to avoid the influence of metal ions on the polysaccharide structure. In step S13, the solution obtained in step S11 and the solution obtained in step S12 are mixed according to a set ratio to ensure that the components are fully mixed without destroying the structure.

[0286] Specifically, the performance of the carrier system was significantly improved by standardizing the preparation process of bio-adjuvants. The selection of chitosan's deacetylation degree (85-95%) and molecular weight (50-150kDa) balanced solubility and bioactivity, thereby enhancing the bioavailability of the drug.

[0287] In some embodiments, in step S2, the buffer is selected as:

[0288] When pH < 7.0, use a mixture of citric acid and disodium hydrogen phosphate;

[0289] When pH ≥ 7.0, use a mixture of potassium dihydrogen phosphate and sodium hydroxide;

[0290] The ionic strength of the buffer solution is 0.1-0.2 mol / L, and when the pH changes by ±0.5, the buffer capacity is not less than 0.02 mol / L.

[0291] Specifically, the citric acid-disodium hydrogen phosphate system (pH < 7.0) has good physiological compatibility, while the potassium dihydrogen phosphate-sodium hydroxide system (pH ≥ 7.0) has strong buffering capacity and chemical stability.

[0292] Specifically, the stability of the active ingredients is ensured through precise control of pH value, ionic strength and buffer capacity. When pH < 7.0, the citric acid-disodium hydrogen phosphate system is used, and when pH ≥ 7.0, the potassium dihydrogen phosphate-sodium hydroxide system is used, which fully considers the stability characteristics of brassinolide under different pH conditions; the setting of ionic strength (0.1-0.2 mol / L) takes into account both osmotic effect and cell compatibility, significantly improving the delivery efficiency and utilization rate of the drug solution.

[0293] In some embodiments, in step S2, the surfactant needs to satisfy:

[0294] There is no precipitation, turbidity or stratification within 48 hours of mixing with brassinolide. The surface tension of the aqueous solution can be reduced to 25-35mN / m, the critical micelle concentration is 0.015-0.025%, and the retention rate of the active ingredient is greater than 85% after 2mm precipitation.

[0295] As an additional explanation, the compatibility stability requirement (no precipitation, turbidity, or stratification for 48 hours) is used to evaluate the physicochemical stability of the drug solution system; surface tension control characterizes the cohesive force of the liquid surface; the critical micelle concentration is the minimum concentration of the surfactant to form micelles; and the resistance to rainwater erosion refers to the retention rate of the active ingredient >85% after 2 mm of precipitation.

[0296] Specifically, the physical and chemical properties of the solution were optimized through a comprehensive evaluation of indicators such as compatibility stability test (48h no precipitation), surface tension control (25-35mN / m), critical micelle concentration (0.015-0.025%) and resistance to rain erosion (retention rate >85%). This not only ensured the stability of the solution, but also significantly improved its wettability and permeability on the plant surface, creating favorable conditions for the full exertion of the efficacy of the drug.

[0297] Example 4: Preparation and performance evaluation test of biological adjuvants for brassinolide preparations

[0298] 1. Purpose of the test:

[0299] The effects of bio-auxiliary preparation process parameters and buffer selection on the performance of brassinolide preparations were studied.

[0300] 2. Test materials:

[0301] (1) Chitosan:

[0302] Specifications: Deacetylation degree 85%, 90%, 95%;

[0303] Molecular weight: 50kDa, 100kDa, 150kDa;

[0304] (2) Water-soluble polysaccharides:

[0305] Carrageenan (analytical grade);

[0306] Gum Arabic (analytical grade);

[0307] Xanthan gum (analytical grade);

[0308] (3) Buffer system:

[0309] Citric acid-disodium hydrogen phosphate system;

[0310] Potassium dihydrogen phosphate-sodium hydroxide system;

[0311] (4) Surfactants:

[0312] Tween-80;

[0313] Span -80;

[0314] Alkyl glycosides;

[0315] Silicone surfactants;

[0316] 3. Test methods:

[0317] (1) Preparation of chitosan solution (step S11):

[0318] Chitosan was added to 1% acetic acid solution;

[0319] Stirring speed: 200r / min;

[0320] Dissolving temperature: 25℃;

[0321] Dissolution time: 4h;

[0322] (2) Preparation of water-soluble polysaccharide solution (step S12):

[0323] The polysaccharide was slowly added to deionized water (conductivity < 0.1 μS / cm);

[0324] Preparation concentration: 2%, 3.5%, 5%;

[0325] Stirring speed: 300r / min;

[0326] Dissolving temperature: 25℃;

[0327] (3) Preparation of the composite (step S13):

[0328] Temperature range: 25-30℃;

[0329] Stirring speed: 200-300r / min;

[0330] Mixing time: 30-40min;

[0331] 4. Buffer selection test:

[0332] (1) When pH < 7.0:

[0333] Citric acid-disodium hydrogen phosphate preparation scheme:

[0334] pH 6.5: 0.1 mol / L citric acid and 0.2 mol / L Na2HPO4 were mixed in a ratio of 1:4.6;

[0335] pH 6.8: 0.1 mol / L citric acid and 0.2 mol / L Na2HPO4 were mixed in a ratio of 1:5.2;

[0336] (2) When pH ≥ 7.0:

[0337] Potassium dihydrogen phosphate-sodium hydroxide preparation scheme:

[0338] pH 7.0: 0.1 mol / L KH2PO4 and 0.1 mol / L NaOH were mixed at a ratio of 1:0.58;

[0339] pH 7.2: 0.1 mol / L KH2PO4 and 0.1 mol / L NaOH were mixed at a ratio of 1:0.68;

[0340] 5. Evaluation of surfactant performance:

[0341] Test results:

[0342] See the table below:

[0343] Table 8 Effects of different surfactants on the properties of brassinolide preparations

[0344]

[0345] Table 9 Effects of different preparation processes on the performance of bio-adjuvant composites

[0346]

[0347] Table 9 Note: CV is the coefficient of variation. The smaller the value, the better the stability.

[0348] 6. Optimization results:

[0349] (1) Optimal preparation process parameters of bio-additives:

[0350] Chitosan: chitosan with a deacetylation degree of 90% and a molecular weight of 100 kDa;

[0351] Water-soluble polysaccharide concentration: 3.5%;

[0352] Composite temperature: 28℃;

[0353] Stirring speed: 250r / min;

[0354] Mixing time: 35min;

[0355] (2) Optimal selection of buffer:

[0356] When pH < 7.0: citric acid-disodium hydrogen phosphate system, ionic strength 0.15 mol / L;

[0357] When pH ≥ 7.0: potassium dihydrogen phosphate-sodium hydroxide system, ionic strength 0.12 mol / L;

[0358] Buffer capacity: not less than 0.02 mol / L when pH changes by ±0.5;

[0359] (3) Optimal choice of surfactant:

[0360] Type: Silicone surfactant;

[0361] Use concentration: 0.02%;

[0362] Performance indicators:

[0363] Surface tension: 26.4mN / m;

[0364] Critical micelle concentration: 0.015%;

[0365] Retention rate after rainwater flushing: 90.2%;

[0366] Thus, it can be seen from Example 4 that:

[0367] (1) This example determined the optimal process parameters for the preparation of bio-adjuvants, ensuring the stability of the carrier system and the drug encapsulation efficiency;

[0368] (2) Through the rational selection of buffer, the stability of the preparation under different pH conditions is controlled, providing a reliable guarantee for practical application;

[0369] (3) The optimized surfactant significantly improved the physical and chemical properties of the solution, enhanced its ability to resist rainwater erosion, and prolonged the duration of its efficacy.

[0370] In some embodiments, in step S31, spraying during the tillering stage includes:

[0371] The first spraying is done when the base of the tillering node begins to swell, and the second spraying is done 10-15 days later;

[0372] Humidify the stem base 2 hours before the first spraying;

[0373] The spraying height is 30-40cm from the leaf surface, and the spraying pressure is 0.15-0.20MPa;

[0374] Add 0.02-0.03% water-soluble boron fertilizer and 0.01-0.02% cytokinin to the spray solution;

[0375] Avoid watering within 4-6 hours after spraying and keep the relative moisture content of the soil at 65-70%.

[0376] Specifically, precise regulation of tillering was achieved through the strategy of spraying in batches (10-15 days apart), pre-humidification pretreatment of the stem base, and the coordinated application of water-soluble boron fertilizer and cytokinin. In particular, micro-environmental conditions conducive to tillering and ear formation were established through the regulation of spraying height, coverage uniformity, and soil moisture, which significantly improved the tillering and ear formation rate and group uniformity.

[0377] In some embodiments, in step S32, the jointing stage spraying process includes:

[0378] The first spraying is carried out when the length of the first internode reaches 0.5-1cm;

[0379] 5-7 days after the first spraying, measure the mechanical strength of the stems, and spray a second time when the mechanical strength of the stems is less than 85% of the standard value;

[0380] When spraying, add 0.02-0.04% potassium fertilizer and 0.01-0.02% salicylic acid to promote stem thickening and lignin formation;

[0381] Ensure that the coverage of the spray on the stem surface reaches more than 90%, the coefficient of variation of droplet distribution uniformity is less than 10%, and the unit area dosage is controlled at 45-60L / hm²;

[0382] No mechanical operation should be carried out within 12 hours after spraying, and the soil moisture should be maintained at 75-80% of the field water holding capacity.

[0383] As an additional note, the jointing stage is a critical period when the stem begins to elongate and grow. During this period, the vascular bundles develop vigorously and the cells elongate actively. The addition of potassium fertilizer can promote the development of stem mechanical tissues and participate in cell wall synthesis and thickening. The addition of salicylic acid can regulate the signal molecules of plant defense responses and induce the expression of genes related to lignin synthesis.

[0384] Specifically, by innovatively introducing the secondary spraying criterion based on the mechanical strength of the stems and through the synergistic application of potassium fertilizer and salicylic acid, the mechanical strength of the stems was significantly enhanced. By controlling the spray coverage and the uniformity of droplet distribution, the uniform distribution of the pesticide solution on the stem surface was ensured, effectively improving the supporting capacity of the stems and reducing the risk of lodging.

[0385] In some embodiments, in step S33, the spraying process during the ear differentiation period includes:

[0386] Select the time when the spikelet length is 1-2 mm for the first spraying;

[0387] 3-5 days after the first spraying, the leaf chlorophyll fluorescence parameter Fv / Fm value is used to determine whether a second spraying is needed. When the Fv / Fm value is lower than 0.75, the second spraying is performed.

[0388] The dosage of liquid per unit area is 45-55L / hm², and 0.02-0.03% putrescine and 0.01-0.02% gibberellin are added at the same time for synergistic promotion;

[0389] During the spraying process, samples were taken every 50m² to check the ear distribution to ensure that the liquid was evenly distributed;

[0390] When spraying on sunny days, use a shade net with a shading rate of 30-40% within 2 hours after spraying, and maintain the relative humidity in the field at 70-80%.

[0391] As an additional note, the spikelet length criterion (1-2 mm) is due to the critical stage of spike primordium differentiation, during which spikelet differentiation is most active; the chlorophyll fluorescence parameter (Fv / Fm value <0.75) can reflect the maximum photochemical efficiency of photosystem II and is a non-destructive method for evaluating the photosynthetic capacity of plants.

[0392] Specifically, the chlorophyll fluorescence parameter (Fv / Fm value) was used as the basis for the second spraying, and the compound application of putrescine and gibberellin significantly promoted the differentiation and development of the ear. At the same time, a quality control system based on sampling and analysis was established, and the application of shading measures effectively improved the uniformity of ear differentiation and fruit setting rate.

[0393] In some embodiments, in step S34, the heading period spraying process includes:

[0394] The first spraying should be done when the flag leaves are fully expanded but the ear has not yet emerged;

[0395] 4-6 days after the first spraying, observe the ear length and spray a second time when the ear length reaches 2-3 cm;

[0396] Add 0.03-0.05% of putrescine and 0.02-0.03% of calcium and magnesium chelating agent to the spray solution;

[0397] Control the spray particle size to 60-90μm, and the amount of liquid used per unit area to 50-60L / hm²;

[0398] When spraying on cloudy days, ensure that the coverage of the liquid on the plant surface reaches more than 85%, the coefficient of variation of the uniformity of the liquid deposition on the ear is less than 15%, and the relative humidity of the canopy is maintained at 75-85% after spraying;

[0399] Monitor the leaf moisture content within 24 hours after spraying, and add additional leaf humidification when it is below 85%.

[0400] As an additional note, the first spraying should be timed when the flag leaf is fully expanded but the ear has not yet emerged, which is the critical period for the transition from reproductive growth to yield formation. During this period, carbon and nitrogen metabolism is most active, and assimilates accumulate in large quantities, making it convenient to measure photosynthetic parameters using a photosynthetic meter and hormone content using HPLC; the ear length monitoring index (2-3cm) is the key criterion for the second spraying, as this stage is sensitive to exogenous hormones; putrescine is an important endogenous growth regulator that can promote cell division and tissue differentiation; calcium and magnesium chelators can provide chelated calcium and magnesium nutrients, participating in cell wall synthesis and signal transduction.

[0401] Specifically, the coordinated regulation of ear development was achieved through the application of putrescine and calcium-magnesium chelators in combination with the split spraying strategy and ear length monitoring. The grain filling efficiency and grain quality were significantly improved by optimizing the particle size and deposition uniformity of the liquid medicine and strengthening the monitoring and regulation of leaf moisture content. Example

[0402] 1. Test materials:

[0403] Test varieties: Jingu 21 (a representative upright plant type variety);

[0404] Test agent: 24-epicrassinolide preparation (prepared according to the optimized formula of Example 4);

[0405] 2. Experimental design:

[0406] Area: 2000m², randomized block design, 3 replications;

[0407] The spraying tests were carried out at tillering stage, jointing stage, ear differentiation stage and heading stage respectively;

[0408] 3. Spraying process parameters and results during tillering period:

[0409] (1) Spraying period: The first spraying is done when the base of the tillering node begins to swell, and the second spraying is done 12 days later;

[0410] (2) Pretreatment of stem base: humidify the stem base 2 hours before spraying, and the water content of the stem base reaches 85% after humidification;

[0411] (3) Spraying height: 35 cm from the leaf surface;

[0412] (4) Solution ratio: add 0.025% water-soluble boron fertilizer and 0.015% cytokinin;

[0413] (5) Soil moisture: Maintain soil relative moisture content at 68%;

[0414] Test results: The effective tillering ear formation rate reached 92%, the vascular bundles of the tillering nodes were well developed, and the tillering angles were highly consistent;

[0415] 4. Spraying process parameters and results during jointing stage:

[0416] (1) Spraying period: The first spraying was carried out when the length of the first internode reached 0.8 cm. The mechanical strength of the stem was measured 6 days later. The second spraying was carried out when it was lower than 85% of the standard value.

[0417] (2) Solution ratio: add 0.03% potassium fertilizer and 0.015% salicylic acid;

[0418] (3) Spray parameters: Stem surface coverage reaches 92%, droplet distribution uniformity coefficient of variation is 8%

[0419] (4) Dosage: 55L / hm² per unit area;

[0420] Test results: The mechanical strength of the stems was significantly improved, the number of vascular bundles increased, and the lignin content increased;

[0421] 5. Spraying process parameters and results during ear differentiation period:

[0422] (1) Spraying period: The first spraying was carried out when the panicle length was 1.5 mm, and the second spraying was carried out 4 days later based on the leaf chlorophyll fluorescence parameter measurement (Fv / Fm value was 0.72);

[0423] (2) Liquid dosage: 50L / hm² per unit area;

[0424] (3) Additives: 0.025% putrescine and 0.015% gibberellin;

[0425] (4) Sampling monitoring: sampling every 100m² to monitor ear differentiation;

[0426] (5) Environmental control: After spraying, use a sunshade net with a shading rate of 35% and maintain the relative humidity in the field at 75%.

[0427] Test results: The uniformity of spikelet differentiation was high, the number of spikelet differentiation increased, and the synchronization of differentiation process was good;

[0428] 6. Spraying process parameters and results at heading stage:

[0429] (1) Spraying period: The first spraying is done when the flag leaf is fully expanded but the ear has not yet emerged. The second spraying is done 5 days later when the ear length reaches 2.5 cm.

[0430] (2) Solution ratio: add 0.04% putrescine and 0.025% calcium and magnesium chelating agent;

[0431] (3) Spray parameters: spray particle size 75 μm, unit area dosage 55 L / hm²;

[0432] (4) Distribution of the liquid medicine: the coverage of the plant surface was 88%, and the coefficient of variation of the deposition uniformity at the ear position was 12%;

[0433] (5) Environmental control: Maintain canopy relative humidity at 80% after spraying;

[0434] (6) Subsequent monitoring: leaf water content is maintained above 88%;

[0435] Test results: The ears are well developed, heading is neat, and the grains are fully filled;

[0436] 7. Conclusion:

[0437] This example verifies the feasibility of spraying process parameters at different growth stages. Through precise spraying at the tillering stage, jointing stage, ear differentiation stage and heading stage, the regulation target of each growth stage is achieved. In particular, a complete set of technical parameter systems has been established in terms of the selection of spraying period, optimization of liquid medicine ratio, and control of environmental conditions, providing technical support for the application of brassinolide in millet production.

[0438] In some embodiments, spraying adjustments under special weather conditions include:

[0439] Rainy weather treatment: When it rains continuously, increase the concentration of the liquid medicine by 15-20%, add 0.02-0.03% of vine extension agent and 0.01-0.02% of pyrrolidone compounds, choose the interval between rains for spraying, and reduce the dosage to 80-85% of the standard dosage;

[0440] High temperature weather treatment: When the temperature is higher than 32℃, spray before the morning dew dries up, reduce the concentration of brassinolide to 70-80% of the standard concentration, add 0.02-0.03% betaine as a protective agent, and use a shade net to shade after spraying, with the shading rate controlled at 40-50%;

[0441] Drought stress treatment: Supplementary spraying is carried out from sunset to after sunset, adding 0.02-0.03% moisturizer and 0.01-0.02% potassium humate. After spraying, cover the ground with straw or biodegradable film in time, increase the number of spraying, and reduce the amount used each time to 60-70% of the standard amount;

[0442] Treatment for windy weather: When the wind speed exceeds 3m / s, suspend spraying. When the wind speed drops below 3m / s, choose to spray row by row in the leeward direction. Re-spray the sprayed area once within 12 hours after the wind speed decreases. During the supplementary spraying, increase the brassinolide concentration by 10-15%.

[0443] Specifically, in rainy weather, vine spreaders are added to improve the spread of the solution on the leaves; in hot weather, betaine is added for osmotic regulation and membrane system protection; in drought stress, the spraying time is selected from sunset to after sunset (such as after 18:00), when plant transpiration is weakened, and humectants can increase the water content of plant tissues, form a water-retaining film, and reduce water loss.

[0444] Specifically, the effect of pesticide application under different climatic conditions has been significantly improved by adjusting the concentration of the liquid medicine, adding specific adjuvants, optimizing the timing and method of spraying, etc. Especially under extreme weather conditions, the scientific supplementary spraying strategy effectively ensures the stability and sustainability of the regulation effect, greatly improving the practicality and adaptability of the technical system.

[0445] Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments, or to make equivalent substitutions for some of the technical features therein. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for spraying exogenous brassinolide on millet, characterized in that: The steps include: Step S1) Recipe preparation: A 24-epi-brassinolide spray solution with a concentration of 0.10-0.25 mg / L is prepared, and the brassinolide raw powder and the biological additive are mixed in a mass ratio of 1:2-1:4, wherein the biological additive is a complex of chitosan and a water-soluble polysaccharide; For erect millet, a composite carrier of chitosan and carrageenan is used, combined with the synergistic effect of pyrrolidone formic acid; for tillering millet, a composite system of chitosan and gum arabic is selected, combined with the osmotic regulation effect of betaine; for hybrid varieties, a carrier system of chitosan and xanthan gum is used, combined with the signal induction effect of amino oligosaccharides; Step S2) Solution preparation: The prepared mixture was diluted with a buffer solution having a pH value of 6.5-7.2, 0.02-0.05% of a surfactant was added, and stirred evenly; Step S3) Spraying during the growth period: S31) Tillering stage: During the peak tillering period, spray at a temperature of 20-25°C with a spraying concentration of 0.10-0.15 mg / L; S32) Jointing stage: Spray when the first internode is elongated, with a spray concentration of 0.12-0.18 mg / L; S33) Ear differentiation stage: Spray at the beginning of spikelet differentiation, with a spraying concentration of 0.14-0.22 mg / L; S34) Heading period: Spray at the heading stage with a concentration of 0.15-0.25 mg / L; Among them, the spraying conditions are selected from 2 hours before sunrise to sunrise or from sunset to 1 hour after sunset, the ambient temperature is controlled at 16-27℃, the relative humidity is controlled at 60-85%, the wind speed is less than 2m / s, and no rainfall is expected within 24 hours.

2. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S1, the formula preparation is selected as follows according to the millet plant type: When the millet is an upright plant type, the concentration of brassinolide is 0.15-0.20 mg / L, chitosan and carrageenan are selected with a mass ratio of 3:1, and 0.03% pyrrolidone formic acid is added; When it is tillering millet, the concentration of brassinolide is 0.10-0.15 mg / L, chitosan and gum arabic are selected with a mass ratio of 2:1, and 0.02% betaine is added; When it is a hybrid millet variety, the brassinolide concentration is 0.20-0.25 mg / L, chitosan and xanthan gum are selected with a mass ratio of 4:1, and 0.04% amino oligosaccharide is added.

3. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S1, the method for preparing the biological additive comprises: Step S11) Preparation of chitosan solution: Chitosan with a deacetylation degree of 85-95% and a molecular weight of 50-150 kDa was selected and dissolved in 1% acetic acid solution; Step S12) Preparation of water-soluble polysaccharide solution: Dissolve the water-soluble polysaccharide in deionized water to prepare a 2-5% aqueous solution; Step S13) Composite preparation: At 25-30° C., the solution obtained in step S11 and the solution obtained in step S12 are mixed according to a set ratio.

4. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S2, the buffer solution is selected as follows: When pH < 7.0, use a mixture of citric acid and disodium hydrogen phosphate; When pH ≥ 7.0, use a mixture of potassium dihydrogen phosphate and sodium hydroxide; The ionic strength of the buffer solution is 0.1-0.2 mol / L, and when the pH changes by ±0.5, the buffer capacity is not less than 0.02 mol / L.

5. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S2, the surfactant must satisfy: There is no precipitation, turbidity or stratification within 48 hours of mixing with brassinolide. The surface tension of the aqueous solution can be reduced to 25-35mN / m, the critical micelle concentration is 0.015-0.025%, and the retention rate of the active ingredient is greater than 85% after 2mm precipitation.

6. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S31, the tillering stage spraying includes: The first spraying is done when the base of the tillering node begins to swell, and the second spraying is done 10-15 days later; Humidify the stem base 2 hours before the first spraying; The spraying height is 30-40cm from the leaf surface, and the spraying pressure is 0.15-0.20MPa; Add 0.02-0.03% water-soluble boron fertilizer and 0.01-0.02% cytokinin to the spray solution; Avoid watering within 4-6 hours after spraying and keep the relative moisture content of the soil at 65-70%.

7. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S32, the jointing stage spraying process includes: The first spraying is carried out when the length of the first internode reaches 0.5-1cm; 5-7 days after the first spraying, measure the mechanical strength of the stems, and spray a second time when the mechanical strength of the stems is less than 85% of the standard value; When spraying, add 0.02-0.04% potassium fertilizer and 0.01-0.02% salicylic acid to promote stem thickening and lignin formation; Ensure that the coverage of the spray on the stem surface reaches more than 90%, the coefficient of variation of droplet distribution uniformity is less than 10%, and the unit area dosage is controlled at 45-60L / hm²; No mechanical operation should be carried out within 12 hours after spraying, and the soil moisture should be maintained at 75-80% of the field water holding capacity.

8. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S33, the spraying process during the ear differentiation period includes: Select the time when the spikelet length is 1-2 mm for the first spraying; 3-5 days after the first spraying, the leaf chlorophyll fluorescence parameter Fv / Fm value is used to determine whether a second spraying is needed. When the Fv / Fm value is lower than 0.75, the second spraying is performed. The dosage of liquid per unit area is 45-55L / hm², and 0.02-0.03% putrescine and 0.01-0.02% gibberellin are added at the same time for synergistic promotion; During the spraying process, samples were taken every 50m² to check the ear distribution to ensure that the liquid was evenly distributed; When spraying on sunny days, use a shade net with a shading rate of 30-40% within 2 hours after spraying, and maintain the relative humidity in the field at 70-80%.

9. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: In step S34, the heading period spraying process includes: The first spraying should be done when the flag leaves are fully expanded but the ear has not yet emerged; 4-6 days after the first spraying, observe the ear length and spray a second time when the ear length reaches 2-3 cm; Add 0.03-0.05% of putrescine and 0.02-0.03% of calcium and magnesium chelating agent to the spray solution; Control the spray particle size to 60-90μm, and the amount of liquid used per unit area to 50-60L / hm²; When spraying on cloudy days, ensure that the coverage of the liquid on the plant surface reaches more than 85%, the coefficient of variation of the uniformity of the liquid deposition on the ear is less than 15%, and the relative humidity of the canopy is maintained at 75-85% after spraying; Monitor the leaf moisture content within 24 hours after spraying, and add additional leaf humidification when it is below 85%.

10. The method for spraying exogenous brassinolide for millet according to claim 1, characterized in that: Adjustments to spraying under special weather conditions include: Rainy weather treatment: When it rains continuously, increase the concentration of the liquid medicine by 15-20%, add 0.02-0.03% of vine extension agent and 0.01-0.02% of pyrrolidone compounds, choose the interval between rains for spraying, and reduce the dosage to 80-85% of the standard dosage; High temperature weather treatment: When the temperature is higher than 32℃, spray before the morning dew dries up, reduce the concentration of brassinolide to 70-80% of the standard concentration, add 0.02-0.03% betaine as a protective agent, and use a shade net to shade after spraying, with the shading rate controlled at 40-50%; Drought stress treatment: Supplementary spraying is carried out from sunset to after sunset, adding 0.02-0.03% moisturizer and 0.01-0.02% potassium humate. After spraying, cover the ground with straw or biodegradable film in time, increase the number of spraying, and reduce the amount used each time to 60-70% of the standard amount; Treatment for windy weather: When the wind speed exceeds 3m / s, suspend spraying. When the wind speed drops below 3m / s, choose to spray row by row in the leeward direction. Re-spray the sprayed area once within 12 hours after the wind speed decreases. During the supplementary spraying, increase the brassinolide concentration by 10-15%.

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