Method for improving drought resistance of plants and related application

Abstract

According to the method for improving the drought resistance of the plants, the plants are pretreated through the tobacco source germination stimulant KAR1, and therefore the drought resistance of the plants is improved. Preferably, the method for improving the drought resistance of the plants specifically comprises the following steps: (1) preparing a KAR1 treating fluid; (2) pretreating plants by adopting a KAR1 treating fluid; the invention also provides an application of the tobacco source germination stimulant KAR1 in improvement of plant drought resistance. According to the method for improving the plant drought resistance and the application of the tobacco source germination stimulant KAR1 in improving the plant drought resistance, the plant drought resistance can be effectively improved, meanwhile, the method is environmentally friendly and safe and harmless to use, the production cost can be reduced, and the method is suitable for large-scale application and popularization.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and more specifically, to the field of plant drought resistance technology, particularly to a method for improving plant drought resistance and related applications. Background Technology

[0002] Plants such as common beans (Phaseolus vulgaris.L.), also known as string beans or kidney beans, belong to the genus Phaseolus in the subfamily Papilionoideae of the family Fabaceae. They are one of the three most important soybean crops globally, accounting for approximately 50% of total edible bean consumption. Common beans are also an important horticultural crop in my country's agricultural restructuring and have become a famous legume vegetable in northern my country and even nationwide. Compared to warm-season beans such as cowpeas and mung beans, common beans are less drought-tolerant. A significant portion of my country's total arable land is low- to medium-yield farmland frequently subjected to drought and other stresses, including major common bean producing areas such as Northeast China, Southwest China, and Xinjiang. Drought stress poses a serious threat to common bean production in my country.

[0003] Applying plant growth regulators for chemical regulation to improve the drought resistance of green beans is an effective solution. Abscisic acid (ABA) can cause rapid stomatal closure, inhibit transpiration, and promote balanced water and fertilizer absorption by plants, making it an important drought-resistant water-retaining agent. However, natural ABA is unstable and difficult to synthesize artificially, resulting in high production costs; some ABA substitutes or water-retaining agents with similar effects have poor environmental friendliness.

[0004] Therefore, it is of practical significance to develop a method that can effectively improve plant drought resistance, is environmentally friendly, safe and harmless to use, and can reduce production costs. Summary of the Invention

[0005] In order to overcome the shortcomings of the prior art, one object of the present invention is to provide a method for improving plant drought resistance, which can effectively improve plant drought resistance, while being environmentally friendly, safe and harmless to use, reducing production costs, and suitable for large-scale promotion and application.

[0006] Another objective of this invention is to provide the application of the tobacco germination stimulant KAR1 in improving plant drought resistance. It can effectively improve plant drought resistance, while being environmentally friendly, safe and harmless to use, reducing production costs, and suitable for large-scale promotion and application.

[0007] To achieve the above objectives, in a first aspect of the present invention, a method for improving plant drought resistance is provided, characterized by pretreating plants with the tobacco germination stimulant KAR1, thereby improving the plant's drought resistance.

[0008] Preferably, the method for improving plant drought resistance specifically includes:

[0009] (1) Prepare KAR1 treatment solution;

[0010] (2) The plants were pretreated with the KAR1 treatment solution.

[0011] More preferably, in step (1), the concentration of the KAR1 treatment solution is 0.5 μM to 1 μM.

[0012] More preferably, step (1) specifically includes:

[0013] (11) Dissolve KAR1 in dimethyl sulfoxide to obtain KAR1 mother liquor;

[0014] (12) Dilute the KAR1 mother liquor with water to obtain the KAR1 treatment solution.

[0015] Furthermore, in step (11), the concentration of the KAR1 mother liquor is 10 mM.

[0016] More preferably, step (2) specifically includes: pretreating the leaves and buds of the plant with the KAR1 treatment solution.

[0017] More preferably, in step (2), the pretreatment is spraying.

[0018] More preferably, in step (2), the plant is a kidney bean.

[0019] Furthermore, in step (2), the green bean is a seedling green bean or a mature green bean.

[0020] In a second aspect of the invention, the application of the tobacco germination stimulant KAR1 in improving plant drought resistance is provided.

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

[0022] 1. The method for improving plant drought resistance of the present invention uses the tobacco germination stimulant KAR1 to pretreat plants, thereby improving the drought resistance of plants. Therefore, it can effectively improve plant drought resistance, and is also environmentally friendly, safe and harmless to use, and can reduce production costs, making it suitable for large-scale promotion and application.

[0023] 2. The application of the tobacco germination stimulant KAR1 of the present invention in improving plant drought resistance can effectively improve plant drought resistance, while being environmentally friendly, safe and harmless to use, reducing production costs, and suitable for large-scale promotion and application.

[0024] These and other objects, features and advantages of the present invention will be fully apparent from the following detailed description and drawings, and can be achieved by the methods, means and combinations thereof specifically pointed out in the specification. Attached Figure Description

[0025] Figure 1 This diagram illustrates the effect of KAR1 treatment on drought resistance in common bean under PEG simulation. A shows the effect of spraying 1 μM KAR1 solution after drought stress on plant phenotype; B shows the effect of normal culture environment and spraying 1 μM KAR1 solution after drought stress on the relative electrical conductivity of common bean plants; C shows the effect of normal culture environment and spraying 1 μM KAR1 solution after drought stress on the maximum photochemical efficiency (Fv / Fm) of photosystem II in common bean leaves. Different lowercase letters indicate differences between treatments reaching a 5% significance level (p<0.05).

[0026] Figure 2 This is a schematic diagram showing the effect of KAR1 treatment on the phenotype of mature beans under progressive drought. A2, B2, and A3 are the plant numbers of beans in the control group, and B4, D2, and D3 are the plant numbers of beans in the KAR1 treatment group.

[0027] Figure 3 This diagram illustrates the effects of KAR1 treatment on the water physiology of the whole bean plant under progressive drought. A) The effect of 0.5 μM KAR1 treatment on the transpiration rate of the whole bean plant under progressive drought; B) The effect of 0.5 μM KAR1 treatment on the diurnal relative turination rate of bean plants under progressive drought; C) The effect of 0.5 μM KAR1 treatment on the midday transpiration rate of bean plants under progressive drought.

[0028] Figure 4 This is a schematic diagram showing the effect of KAR1 treatment on drought resistance in mature beans under natural drought. A shows the effect of 0.5 μM KAR1 treatment on plant phenotype; B shows the effect of spraying 0.5 μM KAR1 treatment on the relative electrical conductivity of bean plants after drought stress. Different lowercase letters indicate that the differences between different treatments reached the 5% significance level (p<0.05). Detailed Implementation

[0029] In order to effectively improve plant drought resistance while being environmentally friendly, safe and harmless to use, and able to reduce production costs, the inventors have conducted extensive research and proposed a method for improving plant drought resistance. This method can effectively improve plant drought resistance while being environmentally friendly, safe and harmless to use, and able to reduce production costs. Based on this, the present invention was completed.

[0030] The method of the present invention for improving plant drought resistance is to pretreat plants with the tobacco germination stimulant KAR1, thereby improving the plant's drought resistance.

[0031] The chemical formula of KAR1, a tobacco germination irritant, is C8H6O3, with a molecular weight of 150.13. Its structural formula is shown below:

[0032] .

[0033] The method for improving plant drought resistance may specifically include any suitable steps; preferably, the method for improving plant drought resistance specifically includes:

[0034] (1) Prepare KAR1 treatment solution;

[0035] (2) The plants were pretreated with the KAR1 treatment solution.

[0036] In step (1), the concentration of the KAR1 treatment solution can be determined as needed. More preferably, in step (1), the concentration of the KAR1 treatment solution is 0.5 μM to 1 μM.

[0037] Step (1) may specifically include any suitable steps, but more preferably, step (1) specifically includes:

[0038] (11) Dissolve KAR1 in dimethyl sulfoxide to obtain KAR1 mother liquor;

[0039] (12) Dilute the KAR1 mother liquor with water to obtain the KAR1 treatment solution.

[0040] In step (11), the concentration of the KAR1 mother liquor can be determined as needed. More specifically, in step (11), the concentration of the KAR1 mother liquor is 10 mM.

[0041] Step (2) may specifically include any suitable steps, but more preferably, step (2) specifically includes: pretreating the leaves and buds of the plant with the KAR1 treatment solution.

[0042] In step (2), the pretreatment can be any suitable pretreatment, and more preferably, in step (2), the pretreatment is spraying.

[0043] In step (2), the plant can be any suitable plant, but more preferably, in step (2), the plant is a common bean.

[0044] In step (2), the green bean can be any green bean at a suitable stage, and more specifically, in step (2), the green bean is a seedling green bean or a mature green bean.

[0045] This invention also provides the application of the tobacco germination stimulant KAR1 in improving plant drought resistance.

[0046] To provide a clearer understanding of the technical content of this invention, the following embodiments are provided for detailed description. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions, such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: ColdSpring Harbor Laboratory Press, 1989), or as recommended by the manufacturer.

[0047] Example 1: Effect of KAR1 on drought resistance of common bean seedlings under PEG simulation

[0048] (1) Plant materials

[0049] Common bean (red flowers and white pods, purchased from Nanjing Green Collar Seed Industry Co., Ltd.)

[0050] (2) Preparation of relevant solutions

[0051] Preparation of 1 / 2 Hoagland nutrient solution: Dissolve 1.74 mM K2SO4, 0.5 mM NH4H2PO4, 2.05 mM MgSO4, and 27.24 µM C in 1 L of water. 10 H 12 The solution consisted of FeN2NaO8, 9.42µM FeSO4, 5.90µM Na2B4O7·10H2O, 7.05µM MnSO4, 0.16µM CuSO4, 0.68µM ZnSO4, 0.8µM (NH4)2SO4, and 2mM Ca(NO3)2·4H2O. The pH was then adjusted to 6.0.

[0052] Preparation of KAR1 stock solution: KAR1 (Shanghai Huicheng Biotechnology Co., Ltd.) and dimethyl sulfoxide (DMSO). DMSO, as an organic solvent, is used to dissolve KAR1, making it easier to mix with other components. This KAR1 stock solution is prepared according to the following steps: Weigh 1.50 mg of KAR1, measure 1 mL of DMSO, and completely dissolve KAR1 in DMSO to obtain a 10 mM KAR1 stock solution.

[0053] Preparation of KAR1 treatment solution and control solution: When applying, dilute the KAR1 stock solution with water to prepare the KAR1 treatment solution. That is, add 100μL of KAR1 stock solution to 1L of aqueous solution containing 200μL of organosilicon (to increase the wetting effect on the leaves), mix the solution evenly, and ensure that no KAR1 is precipitated. The final result is a KAR1 treatment solution containing 1 / 10,000 (volume percentage) DMSO, 2 / 10,000 (volume percentage) organosilicon, and 1μM of active ingredient KAR1.

[0054] The control solution was an aqueous solution containing 1 / 10,000 DMSO and 2 / 10,000 organosilicon.

[0055] (3) Plant material treatment

[0056] When the bean seeds germinate to the point where the hypocotyl elongates but the cotyledons have not yet unfolded, select seedlings of uniform growth, wash them, and transplant them into a hydroponic environment (1 / 2 Hoagland nutrient solution). Change the nutrient solution every 3 days. Once the bean plants have developed three clusters of compound leaves, they are divided into a control group and a KAR1 treatment group, with 8 bean plants in each group. The KAR1 treatment group is sprayed with 1 μM KAR1 solution, while the control group is sprayed with the control solution. Spraying is applied to the upper and lower surfaces of the unfolded leaves and young shoots. Three hours after spraying, a drought treatment is performed by replacing the (1 / 2 Hoagland nutrient solution) with a 1 / 2 Hoagland nutrient solution containing 10% PEG-6000.

[0057] (4) Determination of Fv / Fm

[0058] The maximum photochemical quantum yield (Fv / Fm) of photosystem II was determined using an Imaging-PAM chlorophyll fluorometer. Before testing, bean leaves, after 24 h of drought, were placed in darkness for 30 min to acclimatize. Initial fluorescence (F0) was determined after the measurement light was applied, followed by a 0.8 s saturation pulse (>4000 μmol m). -2 s -1 The maximum fluorescence yield (Fm) of the leaf under darkness was obtained. Fv / Fm = (Fm-F0) / Fm.

[0059] (5) Determination of relative conductivity

[0060] The relative electrolyte permeability (REL) of leaves was determined based on changes in electrical conductivity (EC). Leaves subjected to drought stress for 24 hours were cut into strips, avoiding the main vein. 0.3 g of leaf tissue was weighed and placed in a 50 ml clean centrifuge tube containing 30 ml of deionized water. Background EC (S0) was measured using a conductivity meter. The sample centrifuge tube was placed in a shaker at 28°C and shaken at 200 rpm for 1 hour, after which EC (S1) was measured. Subsequently, the sample was boiled in a water bath for 20 min, and after naturally cooling to room temperature, EC (S2) was measured. The formula for calculating REL (%) is as follows: REL (%) = (S1 - S0) / (S2 - S0) × 100%.

[0061] (6) Experimental results

[0062] Drought stress for 24 hours caused bean plants to wilt and lose water, and the degree of wilting in bean plants treated with KAR1 was less than that in bean plants treated with the control solution. Figure 1 A).

[0063] The results of conductivity measurements showed that before drought treatment, there was no significant difference in the relative conductivity of leaves between the control group and the KAR1 treatment group. After 24 hours of 10% PEG drought stress, the relative conductivity of leaves in both the control group and the KAR1 treatment group increased compared to before drought treatment. The relative conductivity of leaves in the control group was significantly higher than that in the KAR1 treatment group, indicating that after 24 hours of 10% PEG drought stress, the cell membrane permeability of the control group increased compared to the KAR1 treatment group, resulting in greater stress damage. The KAR1 treatment of tobacco germplasm improved the plant's resistance to drought stress. Figure 1 B).

[0064] Fv / Fm is used to measure the primary light energy conversion efficiency of photosystem II in plant leaves. It changes very little under non-stress conditions but decreases significantly under stress conditions. Analysis of Fv / Fm measurements showed that before drought treatment, there was no significant difference in Fv / Fm between the control group and the KAR1 treatment group. After 24 hours of 10% PEG drought stress, the Fv / Fm of the KAR1-treated leaves was not significantly different from that before drought stress, while the Fv / Fm of the control group was significantly lower than before drought stress. This indicates that the plants in the control group suffered some degree of stress damage, while spraying KAR1 could improve the plant's resistance to drought stress. Figure 1 C).

[0065] Example 2: Effects of KAR1 on drought resistance of mature beans under progressive soil drought

[0066] (1) Plant materials

[0067] Common bean (11-P331024004), provided by the Plant Phenotyping and Quality Safety Laboratory of China Jiliang University.

[0068] (2) Test equipment

[0069] Plantarray high-throughput plant physiological characteristics monitoring system (Plant-DiTech, Israel).

[0070] (3) Preparation of treatment solution

[0071] Take 50 μL of the KAR1 mother liquor prepared in Example 1 and add it to 1L of an aqueous solution containing 200 μL of organosilicon. Mix the solution evenly to ensure that no KAR1 is precipitated. Finally, a KAR1 treatment solution containing 0.05% (volume percentage) DMSO, 0.02% (volume percentage) organosilicon, and 0.5 μM of active ingredient KAR1 is obtained.

[0072] The control solution was an aqueous solution containing 0.05% DMSO and 0.02% organosilicon.

[0073] (4) Plant material treatment

[0074] This experiment was conducted at the Plantarray 3.0 high-throughput phenotyping platform (33.62°N, 119.02°E) at the Sino-Israeli Joint Research Center for Plant Physiology in Huai'an City, Jiangsu Province. This platform enables automated, progressive drought treatment. The experimental site was located in a glass greenhouse, with daytime temperatures controlled between 25°C and 30°C and nighttime temperatures controlled between 15°C and 20°C. After calibrating each unit of the Plantarray high-throughput phenotyping platform, they were placed in selected planting pots.

[0075] Healthy seedlings with the first pair of true leaves were transplanted into planting pots, and plastic film was used to cover the pots to prevent water evaporation. Water and fertilizer were precisely controlled through the platform system. Once the seedlings had four leaves and a central bud, 12 units (pots) of healthy, uniformly growing bean plants were selected. Six units were used for the control treatment, and six units were used for the KAR1 treatment. After the system monitoring data stabilized, the experimental treatment began, with a treatment cycle of 20 days. The plants were normally irrigated for 7 days, followed by a gradual drought treatment under platform control from day 8 to day 20, totaling 12 days. One day before the gradual drought (day 7 of the experiment), at 9:00 AM, the plant leaves were treated with either KAR1 or the control solution. The spraying should aim for a uniform, dense distribution of droplets on both sides of the leaf without dripping, with equal application per leaf. Seven days after the first spray (day 15 of the experiment), a second spraying of KAR1 or the control solution was performed. Phenotypic surveys were conducted before the progressive drought treatment, on day 7 of the progressive drought treatment, and on day 12 before rehydration after the progressive drought treatment. Data were recorded against drought severity criteria (Table 1). Water physiological data were processed using the accompanying SPAC analysis software (http: / / spac.plant-ditech.com).

[0076] Table 1. Drought Damage Level Assessment Table

[0077]

[0078] (5) Experimental results

[0079] After 7 days of progressive drought, in the control group, 72% of the bean plants showed grade 2 drought damage, with obvious wilting, water loss, and yellowing of leaves in some areas or throughout the plant. Only 28% of the plants showed grade 1 drought damage, with wilting but no chlorosis in the leaves (Table 2). In the KAR1 treatment group, 33% of the bean plants showed grade 2 drought damage, while the remaining plants showed grade 1 drought damage.

[0080] Table 2. Statistics on Drought Damage Levels of Common Bean Plants

[0081]

[0082] After 12 days of progressive drought, 66% of the plants in the control group showed complete wilt and near-death, with 4 and 2 plants exhibiting grade 2 and grade 1 drought damage, respectively. Compared to the control group, the mortality rate of bean plants treated with KAR1 decreased by 50%. These results indicate that exogenous application of KAR1 can effectively alleviate the stress damage caused by progressive drought to bean plants. Figure 2 ).

[0083] During the 12-day progressive drought, the whole-plant transpiration rate (Tr) of both the control and KAR1 treatment groups exhibited rhythmic variations, with peak values ​​occurring around noon. The maximum diurnal transpiration rate of the control group remained at a high level (around 1.0) from days 1 to 5 of the progressive drought, decreasing to approximately 65% ​​of its original level on day 6. Figure 3 A). Plants in the KAR1 treatment group showed a significantly lower midday transpiration rate compared to the control group from days 1 to 5. Figure 3 A). Further analysis was conducted on the dynamic differences in transpiration rates between the treated and control groups within 8 hours after KAR1 treatment. To reduce interplant differences, the transpiration rate at each treatment time was used as 1 to calculate the relative transpiration rate. The results showed that KAR1 treatment led to a decrease in the relative transpiration rate of the plants, reaching a significant level after 2 hours (A). Figure 3 B). Comparing the midday transpiration rate of different treatments that eliminated the difference in saturated atmospheric pressure (VPD) with changes in soil water content (VWC), it was found that the midday transpiration rate of plants in the KAR1 treatment group was more sensitive to changes in VWC than that of the control group. Figure 3 C). These results indicate that KAR1 treatment may induce a faster plant response to soil drought, close stomata to reduce transpiration in order to maintain plant water homeostasis, and improve drought resistance.

[0084] Example 3: Effects of KAR1 on drought resistance of mature beans under natural soil drought.

[0085] (1) Plant materials

[0086] Common bean (red flowers and white pods, purchased from Nanjing Green Collar Seed Industry Co., Ltd.)

[0087] (2) Preparation of treatment solution

[0088] Take 50 μL of the KAR1 mother liquor prepared in Example 1 and add it to 1L of an aqueous solution containing 200 μL of organosilicon. Mix the solution evenly to ensure that no KAR1 is precipitated. Finally, a KAR1 treatment solution containing 0.05% (volume percentage) DMSO, 0.02% (volume percentage) organosilicon, and 0.5 μM of active ingredient KAR1 is obtained.

[0089] The control solution was an aqueous solution containing 0.05% DMSO and 0.02% organosilicon.

[0090] (3) Plant material treatment

[0091] Two to three seedlings that had grown to two leaves and one bud were transplanted into pots containing approximately 3.9 L of potting soil, covered with plastic film to prevent water evaporation, and watered from the bottom. When the seedlings reached four leaves and one bud, they were divided into two groups: one group was sprayed with KAR1 treatment solution, and the other group was sprayed with control solution. After spraying, irrigation was controlled to simulate natural drought, and soil moisture content was monitored in real time using a soil moisture sensor. Only plants with consistent moisture content trends were compared to determine their drought phenotype.

[0092] (4) Measurement of conductivity

[0093] At the end of the drought experiment, samples of green bean leaves were cut for conductivity measurement, and the specific process was the same as described in Example 1.

[0094] (5) Experimental results

[0095] Comparing the drought phenotypes of the two groups, it was found that the bottom leaves of the control group turned yellow and the plants wilted slightly, while the leaves of the plants in the KAR1 treatment group remained green and did not turn yellow. Figure 4 A).

[0096] After drought treatment, the conductivity of the control group was significantly higher than that of the KAR1 treatment group, indicating that the cell membrane permeability of the control group was increased and the stress damage was greater compared with that of the KAR1 treatment group. The KAR1 treatment of tobacco germplasm improved the plant's resistance to drought stress. Figure 4 B).

[0097] Therefore, by using the tobacco germination stimulant KAR1 to pretreat plants, this invention can effectively improve plant drought resistance, indicating that KAR1 has ABA-like effects. Furthermore, KAR1 is readily available, inexpensive, and environmentally friendly, making it suitable for large-scale promotion and application.

[0098] In summary, the method for improving plant drought resistance of the present invention and the application of tobacco germination stimulant KAR1 in improving plant drought resistance can effectively improve plant drought resistance, while being environmentally friendly, safe and harmless to use, reducing production costs, and suitable for large-scale promotion and application.

[0099] In this specification, the invention has been described with reference to specific embodiments thereof. However, it will be apparent that various modifications and variations can be made without departing from the spirit and scope of the invention. Therefore, the specification and drawings should be considered illustrative rather than restrictive.

Claims

1. A method for improving plant drought resistance, characterized in that, Plants were pretreated with the tobacco germination stimulant KAR1 to improve their drought resistance.

2. The method for improving plant drought resistance according to claim 1, characterized in that, The methods for improving plant drought resistance specifically include: (1) Prepare KAR1 treatment solution; (2) The plants were pretreated with the KAR1 treatment solution.

3. The method for improving plant drought resistance according to claim 2, characterized in that, In step (1), the concentration of the KAR1 treatment solution is 0.5 μM to 1 μM.

4. The method for improving plant drought resistance according to claim 2, characterized in that, Step (1) specifically includes: (11) Dissolve KAR1 in dimethyl sulfoxide to obtain KAR1 mother liquor; (12) Dilute the KAR1 mother liquor with water to obtain the KAR1 treatment solution.

5. The method for improving plant drought resistance according to claim 4, characterized in that, In step (11), the concentration of the KAR1 mother liquor is 10 mM.

6. The method for improving plant drought resistance according to claim 2, characterized in that, Step (2) specifically includes: pretreating the leaves and buds of the plant with the KAR1 treatment solution.

7. The method for improving plant drought resistance according to claim 2, characterized in that, In step (2), the pretreatment is spraying.

8. The method for improving plant drought resistance according to claim 2, characterized in that, In step (2), the plant is a kidney bean.

9. The method for improving plant drought resistance according to claim 8, characterized in that, In step (2), the green bean is a seedling green bean or a mature green bean.

10. Application of KAR1, a tobacco germination stimulant, in improving plant drought resistance.

Images