Method for improving drought tolerance of plants

Abstract

The invention relates to the field of plant cultivation, and discloses a method for improving plant drought tolerance, which comprises the step of pretreating plants with lactose before drought. The method comprises the following steps: by taking a sonchaceae resuscitated plant boea boea, a model plant arabidopsis thaliana, tobacco and crops corn and wheat as research objects, analyzing leaf photosynthesis efficiency, ion leakage and plant survival rate after drought treatment and water supply retreatment on plants subjected to lactose pretreatment; the invention finds that lactose has a positive effect in the aspects of inducing the dehydration resistance of an endiriaceae resuscitation plant boea boea and the drought resistance of model plants arabidopsis thaliana, tobacco and crops corn and wheat, can be applied to inducing the drought resistance of other plants, and is an effective plant drought resistance inducer.

Description

Technical Field

[0001] The invention relates to the field of plant cultivation, and specifically discloses a method for improving drought resistance of plants. Background Art

[0002] With the global shortage of water resources and the increase of extreme climate events, the impact of drought on plant growth, yield and quality is becoming increasingly significant. When organisms are subjected to drought stress or other adverse environmental factors, they will resist the adverse stress through different pathways, including the synthesis and accumulation of drought-resistant related proteins and metabolites.

[0003] Resuscitation plants are considered to be excellent plant materials for studying plant stress resistance mechanisms and exploring stress resistance resources because they can tolerate extremely dry living conditions. There are many plants in the Gesneriaceae family that have the characteristics of dehydration resistance and resuscitation, such as Gesneriaceae ( Boea hygrometrica ), the nutritional tissues of this type of plant have strong drought resistance and recovery capabilities. After growing for about a week in room temperature conditions with nearly dry soil, the relative water content of its leaves drops to about 10%, and photosynthesis basically stops. As long as water is re-applied, the leaves can absorb water and stretch, and return to their appearance and physiological state before dehydration (including the recovery of photosynthesis). Metabolomics studies have found that the content of hormones (including auxin, cytokinin and abscisic acid, etc.), oligosaccharides (including sucrose, maltose, trehalose and lactose, etc.), lipids and phenolic substances in the leaves of Hedera capillaris increases significantly during dehydration.

[0004] Lactose is a disaccharide composed of glucose and galactose, which is widely found in the milk of humans and mammals. A large number of studies have shown that lactose is correlated with the salt tolerance of crops such as rice, soybeans and lentils, and in the resuscitation plant Fanleaf ( Myrothamnus flabellifolius ) and charcoal wood ( Xerophyta schlechteri ). A large amount of lactose was found in the leaves of the revived plant, Lactose, and its content increased significantly during drought stress. Summary of the invention

[0005] The inventors used lactose to treat the recovery plant Heterocarpus fasciatus, the model plant Arabidopsis thaliana, tobacco, and the crops corn and wheat in vitro, and found that the adaptability of the plants pretreated with lactose to drought stress was significantly improved, thereby proposing the present invention.

[0006] The present invention first provides a method for improving drought tolerance of plants, which comprises the step of pre-treating plants with lactose before drought.

[0007] In the above method, the plant is a resuscitation plant, a model plant or a crop.

[0008] In the above method, the drought tolerance of the revived plant may be dehydration tolerance, and the revived plant may be Convolvulus fascicularis.

[0009] In the above method, the model plant may be Arabidopsis thaliana or tobacco.

[0010] In the above method, the crop may be corn or wheat.

[0011] In the above method, the plant is a soil-grown plant, and the lactose pretreatment is carried out by watering with a lactose solution.

[0012] In the above method, the plant is cultivated in a culture medium, and the lactose pretreatment is performed by adding lactose to the culture medium.

[0013] In the above method, the drought tolerance can be embodied as any one or more combinations of the following A1-A (for example, a combination of A1, A2 and A4, or a combination of A1, A3 and A4): A1. Re-watering after drought improves leaf photosynthesis efficiency; A2. Re-watering after drought and dehydration can improve plant recovery rate; A3. Re-watering after drought improves plant survival rate; A4. Re-water after drought and the leaf ion leakage will decrease.

[0014] The present application also provides the use of lactose in preparing a plant drought resistance inducer.

[0015] The active ingredient of the plant drought resistance inducer contains lactose.

[0016] The present invention takes Gesneriaceae resurrection plant Gesneriaceae, model plants Arabidopsis thaliana, tobacco and crops corn and wheat as research objects, and analyzes leaf photosynthesis efficiency, ion leakage and plant survival rate after plants pretreated with lactose are subjected to drought and re-watering treatments, and finds that lactose has a positive effect on inducing dehydration tolerance of Gesneriaceae resurrection plant Gesneriaceae and drought resistance of model plants Arabidopsis thaliana, tobacco and crops corn and wheat. Therefore, lactose can be used to induce drought resistance in other plants, and is an effective inducer of plant drought resistance. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The present invention is based on the plant phenotype, chlorophyll fluorescence and leaf photosynthesis efficiency of the Gesneriaceae resuscitation plant Gesneriaceae cultivated in the soil of Example 1 of the present invention after being pretreated with different concentrations of lactose, rapid drought dehydration treatment and re-watering treatment, as well as the leaf ion leakage and plant recovery rate after re-watering treatment. Figure 1 A is the plant phenotype and chlorophyll fluorescence, Figure 1 B is before and after lactose pretreatment ( Figure 1B (left) and after re-water treatment ( Figure 1 The photosynthesis efficiency of leaves in B (right figure) is Figure 1 C is the leaf ion leakage after re-watering. Figure 1 D is the plant recovery rate after re-watering. ** represents the significant analysis result of P < 0.01, and different lowercase letters represent the significant analysis result of P < 0.05.

[0018] Figure 2 The present invention shows the plant phenotype, chlorophyll fluorescence and leaf photosynthesis efficiency of the aseptically cultivated Gesneriaceae resuscitation plant Gesneriaceae according to Example 2 after lactose pretreatment, rapid drought dehydration and re-watering, as well as the leaf ion leakage and plant recovery rate after re-watering. Figure 2 A is the plant phenotype and chlorophyll fluorescence after lactose pretreatment, rapid drought dehydration and re-watering treatment. Figure 2 B is the photosynthetic efficiency of leaves after lactose pretreatment, rapid drought dehydration and re-watering treatment. Figure 2 C is the leaf ion leakage after re-watering. Figure 2 D is the plant recovery rate after re-watering. * represents the significant analysis result of P < 0.05, ** represents the significant analysis result of P < 0.01, and *** represents the significant analysis result of P < 0.001.

[0019] Figure 3 The plant phenotype photos and leaf photosynthesis efficiency of the model plant Arabidopsis thaliana seedlings pretreated with lactose in Example 3 of the present invention after drought for 8-12 days, as well as the leaf ion leakage and plant survival rate after re-watering. Figure 3 A is a photo of the plant phenotype after re-watering after 8 days of drought, re-watering after 10 days of drought, and re-watering after 12 days of drought. Figure 3 B is before and after lactose pretreatment ( Figure 3 The left figure in B) and the treatments after 8 days of drought, 10 days of drought, and 12 days of drought ( Figure 3 The photosynthesis efficiency of leaves in B (right figure) is Figure 3 C is the leaf ion leakage after 8 days of drought and then watering, 10 days of drought and then watering, and 12 days of drought and then watering. Figure 3 D is the plant recovery rate after re-watering after 8 days of drought, re-watering after 10 days of drought, and re-watering after 12 days of drought. ** represents the significant analysis result of P<0.01, *** represents the significant analysis result of P<0.001.

[0020] Figure 4The plant phenotype photos and leaf photosynthesis efficiency of the model plant tobacco seedlings pretreated with lactose in Example 3 of the present invention after drought for 8-16 days, as well as the leaf ion leakage and plant survival rate after re-watering. Figure 4 A is the phenotype photos of plants after re-watering after 8 days of drought, re-watering after 12 days of drought, and re-watering after 16 days of drought. Figure 4 B is before and after lactose pretreatment ( Figure 4 The left figure in B) and after 8 days of drought and then re-watering, 12 days of drought and then re-watering, and 16 days of drought and then re-watering ( Figure 4 The photosynthesis efficiency of leaves in B (right figure) is Figure 4 C is the leaf ion leakage after 8 days of drought and then watering, 12 days of drought and then watering, and 16 days of drought and then watering. Figure 4 D is the plant recovery rate after 8 days of drought, 12 days of drought, and 16 days of drought. *** indicates a significant difference of P < 0.001.

[0021] Figure 5 These are photos of plant phenotypes, leaf photosynthesis efficiency, leaf ion leakage and plant survival rate of lactose-pretreated corn seedlings in Example 4 of the present invention after two consecutive drought and re-watering treatments. Figure 5 A is a photo of the plant phenotype after two drought and re-watering treatments. Figure 5 B is the photosynthesis efficiency of leaves after two drought and re-watering treatments. Figure 5 C is the leaf ion leakage after two drought and re-watering treatments. Figure 5 D is the survival rate of plants after two drought and re-watering treatments. * represents the significant analysis result of P < 0.05, ** represents the significant analysis result of P < 0.01, and *** represents the significant analysis result of P < 0.001.

[0022] Figure 6 These are photos of plant phenotypes, leaf photosynthesis efficiency, leaf ion leakage and plant survival rate of lactose-pretreated corn seedlings in Example 4 of the present invention after two consecutive drought and re-watering treatments. Figure 6 A is a photo of the plant phenotype after two drought and re-watering treatments. Figure 6 B is the photosynthesis efficiency of leaves after two drought and re-watering treatments. Figure 6 C is the leaf ion leakage after two drought and re-watering treatments. Figure 6 D is the survival rate of plants after two drought and re-watering treatments. * represents the significant analysis result of P < 0.05, ** represents the significant analysis result of P < 0.01, and *** represents the significant analysis result of P < 0.001. DETAILED DESCRIPTION

[0023] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0024] The quantitative tests in the following examples were all repeated three times, and the results were averaged.

[0025] The experimental methods in the following examples are conventional methods unless otherwise specified. The materials, reagents, etc. used in the following examples are commercially available unless otherwise specified.

[0026] The lactose in the following examples is a product of Beijing Solebow Technology Co., Ltd., item number: L8911-500.

[0027] Example 1 Effect of lactose pretreatment on soil cultured resuscitation plants The soil cultivation, lactose pretreatment, dehydration stress treatment and physiological testing procedures of the revived plants are as follows: Soil cultivation: The Gesneriaceae resurrection plant Gesneriaceae (the material is called Boea hygrometrica , recorded in the literature "Run-Ze Sun, Jie Liu, Yuan-Yuan Wang, Xin Deng, DNA methylation-mediated modulation of rapid desiccation tolerance acquisition and dehydration stress memory in the resurrection plant Boea hygrometrica , PLOSGenetics | https: / / doi.org / 10.1371 / journal.pgen.1009549 April 30, 2021”) seeds were sterilized and vernalized and then cultured on a petri dish containing sterile nutrient soil for 3 weeks to form a large number of young shoots. The young shoots were peeled off and transplanted on a petri dish containing sterile nutrient soil for further culture for 3 weeks. The culture conditions were temperature 25±2℃, light 12–16 hours / day, and light intensity 2000–5000 LX.

[0028] After 3 weeks of continuous culture, lactose pretreatment was performed, and two concentrations of lactose aqueous solution were set for treatment, with pure water as the control group: 10 mM lactose pretreatment group: 10 mM lactose aqueous solution was added to the nutrient soil, 5 mL each time, twice a week, for a total of two weeks.

[0029] 50 mM lactose pretreatment group: Add 50 mM lactose aqueous solution to the nutrient soil, 5 mL each time, 2 times / week, for a total of two weeks.

[0030] Control group: Add pure water to the nutrient soil, 5 mL each time, twice a week, for a total of two weeks.

[0031] Three replicates were set up, with 32 plants in each replicate.

[0032] Dehydration stress treatment and re-watering treatment: After lactose irrigation, the plants of each group were removed from the soil, rinsed with sterile water, dried with sterile filter paper, and then moved to a culture dish containing filter paper. They were subjected to rapid air drought dehydration for two days at a temperature of 25±2℃ and a relative humidity of 60% (i.e., dehydration stress treatment), and then water was added to the filter paper to resume cultivation for 3 days (i.e., re-watering treatment).

[0033] Physiological testing: The modulated chlorophyll fluorescence imaging system was used to detect the leaf photosynthesis efficiency of the Lactobacillus thunbergii seedlings before and after lactose pretreatment, after rapid dehydration treatment and after re-watering treatment. The conductivity meter was used to detect the leaf ion leakage of the Lactobacillus thunbergii seedlings after re-watering treatment, and the plant recovery rate of the Lactobacillus thunbergii seedlings after re-watering treatment was calculated.

[0034] In this example, the soil-cultivated Gesneriaceae resuscitation plant, Gesneriaceae, was used as the research object. Different concentrations of lactose aqueous solution were used to irrigate the soil of Gesneriaceae seedlings. The changes in the photosynthesis efficiency of Gesneriaceae leaves before and after the treatment were analyzed. The photosynthesis efficiency of the leaves, ion leakage detection and plant recovery rate of the isolated plants after rapid drought dehydration and re-watering were also analyzed. The results are shown in Table 1. Figure 1 It was found that low-concentration lactose pretreatment had no significant effect on the photosynthesis efficiency of the leaves of Heliconia seedlings. The leaf photosynthesis efficiency and plant recovery rate of Heliconia seedlings pretreated with low and high concentrations of lactose after rapid drought dehydration and re-watering were significantly improved, and the leaf ion leakage was significantly reduced, proving that lactose pretreatment can significantly improve the dehydration resistance of Heliconia seedlings cultivated in soil.

[0035] Example 2 Effect of lactose pretreatment on tissue culture resuscitation plants The sterile culture, lactose pretreatment, dehydration stress treatment and physiological testing procedures of the revived plants are as follows: Aseptic culture: After the seeds of the Gesneriaceae resurrection plant Gesneriaceae were sterilized and vernalized, they were placed on 1 / 2 MS medium containing 10 g / L sucrose for 3 weeks to form a large number of young shoots. The young shoots were peeled off and transplanted on 1 / 2 MS medium containing 10 g / L sucrose for another 3 weeks. The culture conditions were 25±2℃, 12–16 hours / day of light, and 2000–5000 LX of light intensity.

[0036] After 3 weeks of continuous culture, lactose pretreatment was performed: Lactose pretreatment group: 1.5-month-old sterile seedlings of Lophatherum volutum were transplanted and cultured on 1 / 2 MS medium containing 5 g / L lactose and 5 g / L sucrose for 2 weeks.

[0037] Control group: 1.5-month-old sterile seedlings of L. volvacea were transplanted on 1 / 2 MS medium containing 10 g / L sucrose and cultured for 2 weeks.

[0038] Six replicates were set up, with 30 plants per group in each replicate.

[0039] The culture conditions were a temperature of 25 ± 2 °C, a light intensity of 12–16 h / day, and a light intensity of 2000–5000 LX.

[0040] Dehydration stress treatment and re-watering treatment: After lactose irrigation, the plants of each group were removed from the soil, rinsed with sterile water, dried with sterile filter paper, and then moved to a culture dish containing filter paper. They were subjected to rapid air drought dehydration for two days at a temperature of 25±2℃ and a relative humidity of 60% (i.e., dehydration stress treatment), and then water was added to the filter paper to resume cultivation for 3 days (i.e., re-watering treatment).

[0041] Physiological testing: The modulated chlorophyll fluorescence imaging system was used to detect the leaf photosynthesis efficiency of the Lactose pretreatment, air rapid dehydration and re-watering treatment of the Lactose seedlings. The conductivity meter was used to detect the leaf ion leakage of the Lactose seedlings after re-watering, and the plant recovery rate of the Lactose seedlings after re-watering was calculated.

[0042] In this example, the aseptically cultivated resuscitation plant Heterocarpus volutus was used as the research object. By adding lactose to the culture medium, the aseptic seedlings of Heterocarpus volutus were treated in vitro to analyze the effect of lactose on the photosynthesis efficiency of the leaves of Heterocarpus volutus seedlings. The photosynthesis efficiency, ion leakage detection and recovery rate of the leaves of the isolated plants after rapid drought dehydration and re-watering were analyzed. The results are shown in Figure 2 It was found that adding lactose to the culture medium had no significant effect on the photosynthetic efficiency of the leaves of the Hesperus rotundus seedlings. The leaves of the Hesperus rotundus seedlings pretreated with lactose had significantly improved photosynthetic efficiency and recovery rate after rapid drought dehydration and re-watering, and the leaf ion leakage was significantly reduced, proving that lactose pretreatment can significantly improve the dehydration resistance of Hesperus rotundus sterile seedlings.

[0043] Example 3 Effect of lactose pretreatment on soil culture model plants Model plant cultivation, lactose pretreatment, drought stress treatment and physiological testing process: The seeds of the model plants Arabidopsis thaliana and tobacco were sterilized and vernalized and cultured on 1 / 2 MS medium containing 10 g / L sucrose for 3 weeks to form a large number of young shoots. The young shoots were peeled off and transplanted on culture dishes containing sterile nutrient soil for 3 weeks. The culture conditions were temperature 25±2℃, light 12–16 hours / day, and light intensity 2000–5000 LX.

[0044] After 3 weeks of cultivation, lactose pretreatment was performed. Each plant was treated with 10 mM lactose aqueous solution, and pure water was used as the control group: Arabidopsis lactose pretreatment group: 10 mM lactose aqueous solution was irrigated into the nutrient soil, 1 mL each time, twice a week, for a total of two weeks.

[0045] Arabidopsis control group: irrigate the nutrient soil with pure water, 1 mL each time, twice a week, for a total of two weeks.

[0046] Tobacco lactose pretreatment group: 10 mM lactose aqueous solution was irrigated into the nutrient soil, 1 mL each time, twice a week, for a total of two weeks.

[0047] Tobacco control group: irrigate the nutrient soil with pure water, 1 mL each time, twice a week, for a total of two weeks.

[0048] Drought treatment and re-watering treatment: After lactose pretreatment, drought treatment was carried out, and the Arabidopsis control group, tobacco lactose pretreatment group and tobacco control group were each subjected to three water-stopping treatments, among which Arabidopsis stopped watering for 8 days, 10 days and 12 days, respectively, and tobacco stopped watering for 8 days, 12 days and 16 days, respectively. Then water was re-watered for 3 days.

[0049] Three replicates were set for the number of days of water withdrawal treatment in each group, and 16 plants were treated in each replicate and group.

[0050] Physiological testing: A handheld chlorophyll fluorescence meter was used to detect the leaf photosynthesis efficiency of Arabidopsis and tobacco seedlings before and after lactose pretreatment, drought treatment (Arabidopsis: 8 days, 10 days, 12 days, tobacco: 8 days, 12 days, 16 days) and re-watering after various drought treatments. A conductivity meter was used to detect leaf ion leakage of Arabidopsis and tobacco seedlings after re-watering, and the plant survival rate of Arabidopsis and tobacco seedlings after re-watering was calculated.

[0051] In this example, Arabidopsis thaliana and tobacco, model plants grown in soil, were used as research objects. Lactose aqueous solution was used to irrigate the soil of Arabidopsis thaliana and tobacco seedlings, and the photosynthetic efficiency, ion leakage and plant survival rate of leaves were analyzed after drought and re-watering of the plants. The results of Arabidopsis thaliana are shown in Figure 3 , the tobacco results data are shown in Figure 4It was found that the leaf photosynthesis efficiency and plant survival rate of Arabidopsis thaliana pretreated with lactose were significantly improved and the leaf ion leakage was significantly reduced after drought and re-watering treatments. The leaf ion leakage of tobacco seedlings pretreated with lactose was significantly reduced after drought and re-watering treatments, proving that lactose pretreatment can significantly improve the drought resistance of model plants.

[0052] Example 4 Effect of lactose pretreatment on soil-grown crops Crop cultivation, lactose pretreatment, drought stress treatment and physiological testing process: The seeds of the crops corn and wheat were sterilized and vernalized and then placed on culture dishes containing sterile nutrient soil for 4 weeks to form seedlings. The culture conditions were temperature 25±2℃, light 12–16 hours / day, and light intensity 2000–5000 LX.

[0053] After 4 weeks of cultivation, lactose pretreatment was performed. Each plant was treated with 10 mM lactose aqueous solution, and pure water was used as the control group: Corn lactose pretreatment group: 10 mM lactose aqueous solution was irrigated into the nutrient soil, 5 mL each time, twice a week, for a total of two weeks.

[0054] Corn control group: irrigate the nutrient soil with pure water, 5 mL each time, twice a week, for a total of two weeks.

[0055] Wheat lactose pretreatment group: 10 mM lactose aqueous solution was irrigated into the nutrient soil, 5 mL each time, twice a week, for a total of two weeks.

[0056] Wheat control group: irrigate the nutrient soil with pure water, 5 mL each time, twice a week, for a total of two weeks.

[0057] Three replicates were set up, with 16 plants per group per replicate for each plant.

[0058] Drought treatment and re-watering treatment: After the lactose pre-treatment, drought treatment (withdrawal of water for 16 days) and re-watering for 3 days were carried out, and then drought treatment (withdrawal of water for 16 days) and re-watering for 3 days were repeated.

[0059] Physiological testing: A handheld chlorophyll fluorescence meter was used to detect the photosynthetic efficiency of the leaves of corn and wheat seedlings after two re-watering treatments. A conductivity meter was used to detect the leaf ion leakage of corn and wheat seedlings after two re-watering treatments. The survival rate of the corn and wheat seedlings after two re-watering treatments was also counted.

[0060] In this example, corn and wheat, crops grown in soil, were used as research objects. Soil irrigation was performed on corn and wheat seedlings using lactose aqueous solution. Leaf photosynthesis efficiency, ion leakage and plant survival rate were analyzed after drought and re-watering of the plants. The test results of corn were shown in Table 1. Figure 5 The results of wheat determination are shown in Figure 6 It was found that the leaf photosynthesis efficiency and plant survival rate of corn and wheat pretreated with lactose were significantly improved, while the leaf ion leakage was significantly reduced after two consecutive drought and re-watering treatments, proving that lactose pretreatment can significantly improve the drought resistance of model plants and crops.

[0061] In summary, lactose plays a positive role in inducing dehydration tolerance in the Gesneriaceae resurrection plant Gesneriaceae and drought resistance in model plants Arabidopsis thaliana, tobacco, and crops corn and wheat. Therefore, it can be used to induce drought resistance in other plants and is an effective inducer of plant drought resistance.

[0062] The present invention has been described in detail above. It will be apparent to those skilled in the art that the present invention may be implemented in a wide range under equivalent parameters, concentrations and conditions without departing from the spirit and scope of the present invention and without the need for unnecessary experimentation. Although the present invention provides specific embodiments, it should be understood that further improvements may be made to the present invention. In short, according to the principles of the present invention, this application intends to include any changes, uses or improvements to the present invention, including changes made by conventional techniques known in the art that depart from the scope disclosed in this application. Applications of some of the basic features may be made within the scope of the following appended claims.

Claims

1. A method for improving drought tolerance of plants, characterized in that: The method comprises the step of pre-treating the plant with lactose prior to drought.

2. The method according to claim 1, characterized in that The plant is a resurrection plant, a model plant or a crop.

3. The method according to claim 2, characterized in that The plant is a resuscitation plant, and the drought tolerance is dehydration tolerance.

4. The method according to claim 2, characterized in that The resuscitation plant may be Convolvulus fasciata.

5. The method according to claim 2, characterized in that The model plant is Arabidopsis thaliana or tobacco.

6. The method according to claim 2, characterized in that The crop is corn or wheat.

7. The method according to claim 2, characterized in that The plants are soil-grown plants, and the lactose pretreatment is carried out by watering with lactose solution.

8. The method according to claim 2, characterized in that The plant is cultivated in a culture medium, and the lactose pretreatment is carried out by adding lactose to the culture medium.

9. The method according to any one of claims 1 to 8, characterized in that: The drought tolerance is embodied in any one or more combinations of the following A1-A: A1. Re-watering after drought improves leaf photosynthesis efficiency; A2. Re-watering after drought and dehydration can improve plant recovery rate; A3. Re-watering after drought improves plant survival rate; A4. Re-water after drought and the leaf ion leakage will decrease.

10. Application of lactose in the preparation of plant drought resistance inducers.

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