A method to alleviate cadmium toxicity during buckwheat germination and seedling stages
The synergistic treatment of melatonin and cobalt chloride solved the problem of cadmium toxicity in buckwheat germination and seedling stage, improved the germination rate of buckwheat seeds and the growth recovery ability of seedlings, reduced cadmium content, and effectively alleviated cadmium stress.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- XICHANG COLLEGE
- Filing Date
- 2023-10-09
- Publication Date
- 2026-06-30
Smart Images
Figure CN117397418B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of biotechnology, specifically to a method for mitigating cadmium toxicity during the germination and seedling stages of buckwheat. Background Technology
[0002] Tartary buckwheat (Fagopyrum tataricum (L.) Gaertn.) is one of my country's important dual-purpose grain crops, valued for both its medicinal and culinary uses. It is known as the "King of Grains" and is a traditional staple food for the Yi people of Liangshan Prefecture. Seed germination plays a crucial role in the cultivation of tartary buckwheat, significantly influencing seedling emergence and yield. The germination of tartary buckwheat seeds is determined by both their genetic characteristics and external environmental factors.
[0003] Heavy metal pollution generally refers to pollution caused by metals or their compounds with a specific gravity of 4 or higher (mainly including copper, zinc, chromium, lead, mercury, nickel, cobalt, etc.) in the environment. Among various heavy metal pollutants, cadmium pollution covers the widest area, and various cadmium pollution incidents have emerged frequently in recent years. Cadmium is highly toxic, mainly originating from wastewater discharged from electroplating, mining, smelting, fuel, battery, and chemical industries. When absorbed in large quantities by plants, it can produce various physiological toxic reactions, leading to decreased root vitality, tissue chlorosis, stunted growth, and reduced dry matter yield. The accumulation of cadmium in plants can damage cell structure, reduce root vitality, slow seedling growth, and cause leaves to curl or even turn yellow, inhibiting plant growth and reducing the absorption of water and nutrients. In addition, it can affect plant physiological activities, mainly manifested in the impact on photosynthesis, transpiration, respiration, protective enzyme activity, and cell membrane permeability, ultimately reducing crop yield and quality. Furthermore, cadmium can also enter the human body through the food chain, threatening human health. Therefore, preventing or reducing the absorption of cadmium by crops is an important way to prevent cadmium from being ingested by the human body.
[0004] Melatonin (MT) is a small molecule compound ubiquitous in organisms. As a novel plant hormone, it participates in various physiological processes and acts as a regulator of both abiotic and biotic stresses. Studies have found that melatonin promotes the biosynthesis of glutathione and phytohemagglutinin, thereby isolating cadmium in vacuoles and enhancing cadmium tolerance in tomatoes. Exogenous application of melatonin effectively improves the growth of cotton seedlings under cadmium stress, enhancing their photosynthetic capacity and root development, thus improving cotton's cadmium tolerance. It can also alleviate oxidative stress caused by cadmium toxicity, enhance photosynthetic efficiency and cellular water uptake, and improve tobacco's cadmium tolerance. Melatonin promotes the biosynthesis of downstream sulfur metabolites cysteine and glutathione, thereby improving plant cadmium tolerance by regulating chelating capacity and redox balance.
[0005] Although many studies suggest that melatonin can effectively reduce the absorption and accumulation of cadmium in plants and decrease the translocation of cadmium from the roots to the above-ground parts, the effect of treating buckwheat seeds with melatonin alone is not ideal in practical applications. Therefore, it is necessary to provide a method that can effectively alleviate cadmium toxicity in buckwheat during germination and seedling stages, thereby providing an effective means to understand the cadmium resistance mechanism of buckwheat, improve its stress resistance, and control heavy metal pollution. Summary of the Invention
[0006] In view of the above problems, the purpose of this invention is to overcome the defects in the prior art and provide a method to alleviate cadmium toxicity during the germination and seedling stages of buckwheat.
[0007] This invention is achieved through the following technical solution:
[0008] A method for mitigating cadmium toxicity during the germination and seedling stages of buckwheat, comprising treating plant seeds with a melatonin reagent before germination.
[0009] Optionally, the method includes using a melatonin reagent that is a mixture of melatonin and cobalt chloride.
[0010] Optionally, the method includes a melatonin concentration of 100-500 μmol / L in the melatonin reagent.
[0011] Optionally, the method includes a cobalt chloride concentration of 1-5 μmol / L in the melatonin reagent.
[0012] Optionally, the method includes contacting buckwheat seeds with a melatonin solution under the following conditions: soaking the buckwheat seeds in the melatonin solution for 12-24 hours in the dark at a temperature of 20-28°C.
[0013] The method provided by this invention can effectively improve the germination potential and germination rate of buckwheat seeds under cadmium stress, enabling seeds to germinate uniformly and rapidly. It can also alleviate the inhibitory effect of cadmium stress on the growth of buckwheat seedlings, restoring seedling growth to normal levels. Simultaneously, it reduces the cadmium content in the radicle and plumule of buckwheat. Attached Figure Description
[0014] Figure 1 The effect of different concentrations of melatonin reagent on the germination rate of buckwheat seeds.
[0015] Figure 2 The effect of different concentrations of melatonin reagent on the radicle length after germination of buckwheat seeds.
[0016] Figure 3 The effect of different concentrations of melatonin reagent on the length of the embryo after germination of buckwheat seeds.
[0017] Figure 4The effect of different concentrations of melatonin reagent on the radicle / hypocotyl weight after germination of buckwheat seeds.
[0018] Figure 5 The effect of different concentrations of melatonin reagent on cadmium accumulation after germination of buckwheat seeds.
[0019] Figure 6 To investigate the effects of different concentrations of melatonin reagent on the growth of buckwheat seedlings caused by cadmium toxicity. Detailed Implementation
[0020] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer shall apply. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased commercially.
[0021] The inventors of this invention have conducted extensive and in-depth research on the plant growth regulation mechanism of melatonin, especially on how to improve the cadmium resistance of tartary buckwheat. They have finally proposed a method of using melatonin and cobalt chloride to synergistically resist cadmium. This method has a significant synergistic effect compared to using melatonin alone, and can effectively alleviate cadmium toxicity in tartary buckwheat during germination and seedling stages.
[0022] Specifically, the present invention provides a method for promoting the germination of buckwheat seeds under cadmium stress, wherein the method includes treating the buckwheat seeds with a melatonin reagent before germination.
[0023] In this invention, germination refers to the sprouting of seeds, and the germination site can be a suitable place for seed germination, such as a culture medium, filter paper, or soil.
[0024] In the method provided by the present invention, after the treatment, the buckwheat seeds are washed with distilled water and then air-dried. The purpose of air-drying is to restore the buckwheat seeds to their original weight before treatment.
[0025] In this invention, the contact methods include, but are not limited to, soaking and spraying. Preferably, the contact method is to soak the seeds in the melatonin reagent solution. When using the soaking method to treat the seeds, the amount of melatonin solution should be sufficient to submerge the seeds.
[0026] In this invention, the contact conditions are as follows: the plant seeds are soaked in a melatonin solution for 12-24 hours under dark conditions. The soaking temperature is 20-28°C.
[0027] In the method provided by this invention, after the buckwheat seeds are treated as described above, the seeds can still germinate normally under severe cadmium stress conditions (according to the standard given in GB15618).
[0028] The features and performance of the present invention will be further described in detail below with reference to embodiments.
[0029] Example 1: Preparation of melatonin reagent
[0030] The main component is: 98% melatonin (C 13 H 16 N2O2, Shanghai Maclean Biochemical Technology Co., Ltd.), cobalt chloride (CoCl) 2， Aladdin), anhydrous ethanol, and water.
[0031] The preparation steps are as follows: Weigh 0.1161g of melatonin, dissolve it in 3mL of anhydrous ethanol, and then bring the volume to 50mL to obtain a 10mM melatonin stock solution. Weigh 6.5mg of cobalt chloride, dissolve it in 5mL of anhydrous ethanol, and then bring the volume to 50mL to obtain a 1mM cobalt chloride stock solution. Prepare the melatonin reagent as shown in Table 1.
[0032] Table 1
[0033] Grouping melatonin concentration μmol / L Cobalt chloride concentration (μmol / L) 1 300 0 2 100 1 3 150 2 4 200 3 5 300 4 6 500 5 7 1000 5
[0034] Example 2 Seed Treatment
[0035] Select approximately 1700 uniformly sized, plump, and pest-free buckwheat seeds and place them in a 50mL centrifuge tube. First, disinfect with 75% alcohol for 1 minute, then disinfect with 0.1% mercuric chloride for 20 minutes, shaking the centrifuge tube continuously during this process to ensure thorough disinfection of the seeds. Finally, rinse with deionized water 8-10 times to remove the mercuric chloride.
[0036] The experiment included one positive control (CK) and seven different concentrations of melatonin as experimental groups. The procedure was as follows: the positive control group was treated by soaking the sterilized seeds in pure water, and the experimental groups were treated by soaking the sterilized seeds in the seven groups of melatonin reagents prepared in Example 1. The buckwheat seeds were soaked in the solution for 12-24 hours in the dark, and then the seeds were taken out, washed, and dried.
[0037] The treated buckwheat seeds were placed in 120mm diameter petri dishes lined with clean paper (moistened with the solution of the above concentration), with 70 seeds per dish. 7mL of the above solution of different concentrations was added sequentially, with each concentration treatment repeated three times. The experimental group was treated with 250μM cadmium chloride solution to create a cadmium stress environment. The petri dishes were then placed in a constant temperature incubator at 25℃ and 60% humidity. Distilled water was added daily at a fixed time using a weighing method to replenish the evaporated water, and the experiment was repeated three times. Germination was defined as the emergence of the radicle from the seed coat. The number of germinated seeds and the peak germination period were continuously observed and recorded. The germination experiment ended when no new seeds germinated for four consecutive days. Germination rate, radicle length, plumule length, biomass, and cadmium content were measured.
[0038] The results are as follows Figure 1-5 As shown, the synergistic treatment of melatonin and cobalt chloride improved the germination efficiency of buckwheat seeds compared to the use of melatonin alone. Figure 1 The synergistic treatment of melatonin and cobalt chloride significantly increased the length of the radicle in germinating buckwheat seeds compared to melatonin alone. Figure 2 ). Figure 5 The results show that the cadmium content in the germ and bacon of buckwheat seeds was significantly reduced in the synergistic group compared with the group using only the cadmium and bacon, thus alleviating the toxic effects of cadmium stress on the germ and radicle of buckwheat seeds after germination and promoting the accumulation of dry matter.
[0039] Seeds that germinate under normal conditions (pure water) are transplanted into 1 / 2 Hogland nutrient solution after the taproot grows to about 3cm. Six treatments are set up: a positive control group (1 / 2 Hogland nutrient solution), experimental groups with different concentrations of melatonin (1 / 2 Hogland nutrient solution + component 1-5 + 100μM cadmium chloride), and cultured for 7 days. Then, the root length, aboveground dry weight and underground dry weight are measured.
[0040] The results are as follows Figure 6 As shown, the synergistic treatment with melatonin and cobalt chloride has a significant effect on alleviating the inhibitory effect of cadmium stress on the growth of buckwheat seedlings, and the root system and aboveground parts grow better than those under cadmium stress.
[0041] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.
Claims
1. A method for relieving the sprouting of tartary buckwheat and the cadmium toxicity in seedling stage, characterized in that, The method is to treat the plant seeds with a melatonin reagent before germination, the melatonin reagent is a mixed reagent of melatonin and cobalt chloride, the concentration of melatonin in the melatonin reagent is 100-500 μmol / L, and the concentration of cobalt chloride in the melatonin reagent is 1-5 μmol / L.
2. The method of claim 1, wherein, The method comprises contacting tartary buckwheat seeds with the melatonin reagent.
3. The method of claim 2, wherein, The contacting condition is that the tartary buckwheat seeds are soaked in the melatonin reagent for 12-24 h under dark conditions.
4. The method of claim 3, wherein, The temperature condition of the soaking is 20-28 DEG C.