A magnesium-doped carbon dot and its application and method for promoting tobacco seed germination

Abstract

The present invention provides a magnesium-doped carbon dot and its application and a method for promoting tobacco seed germination. The preparation method of the magnesium-doped carbon dot comprises the following steps: 1) mixing tobacco powder and reduced glutathione in a mass ratio of 1:(1.5-4), dissolving the mixture in formamide, and ultrasonically dispersing the mixture to obtain a reaction solution A; dissolving magnesium hydroxide in formamide at a mass ratio of 1.2-1.4 times that of the tobacco powder, and ultrasonically dispersing the mixture to obtain a reaction solution B, which is mixed with the reaction solution A and placed in an autoclave for reaction; 2) after the reaction is completed, cooling, filtering, transferring the obtained filtrate to a dialysis bag, and freeze-drying the dialyzed solution to obtain the magnesium-doped carbon dot. The present invention belongs to the field of bio-agricultural technology, and the magnesium-doped carbon dot provided can effectively promote the germination of tobacco seeds and is conducive to improving the level of strong seedling cultivation.

Description

Technical Field

[0001] The present invention belongs to the field of bio-agricultural technology, and in particular relates to magnesium-doped carbon dots, applications thereof, and a method for promoting tobacco seed germination. Background Art

[0002] Tobacco is an important economic crop in my country, ranking first in the world in terms of cultivated area and yield. Tobacco is a thermophilic crop with high requirements for light and water, but it is afraid of waterlogging. Seeds are important agricultural production materials. The quality of seeds is mainly reflected by seed vitality, and is usually comprehensively evaluated based on indicators such as seed germination rate and germination index. Seed germination is the beginning of the plant growth cycle and a critical period in the plant's life history. During this stage, the plant is easily affected by environmental factors such as moisture, temperature, and light. Guizhou Province is located in a low-temperature and low-light area with many karst mountains and is prone to drought. The long-term low temperature, low light, and drought climate in the early stage of seedling cultivation have become important constraints on tobacco seed germination and the cultivation of strong seedlings, resulting in low seed germination rate, weak tobacco seedlings, and difficulty in timely seedling formation.

[0003] Carbon dots (CDs) are a new type of nanocarbon material composed of dispersed spherical carbon particles. They are extremely small (below 10 nm) and have fluorescent properties. They also have the advantages of simple preparation, low toxicity, biocompatibility, and ease of functionalization. Carbon dot nanomaterials have shown great application potential in fields such as agriculture and materials. Chinese patent application CN113115605 A discloses a method for promoting spinach seed germination using carbon dots. The carbon dots are prepared into a carbon dot aqueous solution with a concentration of 1-50 mg / L and then applied to spinach seeds. The seeds are then incubated in the dark, which increases the expression of aquaporin genes, promotes spinach seed germination, and improves the germination rate. Chinese patent application CN 113201330A discloses magnesium-nitrogen-doped carbon dots, a preparation method thereof, and use thereof in improving plant photosynthesis. The preparation steps of the magnesium-nitrogen-doped carbon dots include: dissolving citric acid, ethanolamine, and magnesium hydroxide in ultrapure water, pouring the resulting mixture into a reactor after ultrasonic treatment and reacting at 200°C for 6 hours; cooling the reaction temperature to room temperature, filtering the product using a filter with a pore size of 0.22 μm, and then dialyzing it for 12 hours using a dialysis bag with a molecular weight cutoff of 100-500, followed by freeze-drying to obtain the magnesium-nitrogen-doped carbon dots. The invention discloses that rice seeds are cultured in the dark during the germination stage and treated with different concentrations of magnesium-nitrogen-doped carbon dots by foliar spraying to accelerate the metabolic rate of chlorophyll, thereby improving its activity and light absorption capacity.

[0004] Tobacco seed germination requires high levels of light and temperature. Appropriate light conditions promote germination, while excessive or insufficient light can negatively impact seed germination. Tobacco seed germination occurs at a specific temperature, with the optimum temperature being around 25-28°C and the minimum being 7.5-10.0°C. Tobacco seed germination conditions differ significantly from those of spinach and rice seeds. To improve tobacco seed germination and seedling vigor, providing a novel magnesium-doped carbon dot, its application, and methods for promoting tobacco seed germination are of great significance. Summary of the Invention

[0005] To solve the problems existing in the prior art, the present invention studies the effects of carbon dot nanomaterials on naked MS Yunyan 87 seeds under different environmental stresses from three perspectives: weak light stress, PEG-simulated drought stress, and low temperature stress. Furthermore, a magnesium-doped carbon dot and its application and a method for promoting tobacco seed germination are provided, which can effectively promote tobacco seed germination and help improve the level of seedling cultivation.

[0006] The objects of the present invention will be further illustrated by the following detailed description.

[0007] In one aspect, the present invention provides a magnesium-doped carbon dot, the preparation method of which comprises the following steps:

[0008] 1) Tobacco powder and reduced glutathione are mixed at a mass ratio of 1:(1.5-4), dissolved in formamide, and ultrasonically dispersed to obtain reaction solution A; magnesium hydroxide is dissolved in formamide at a mass ratio of 1.2-1.4 times that of the tobacco powder, and ultrasonically dispersed to obtain reaction solution B, which is mixed with reaction solution A and then placed in a high-pressure reactor for reaction;

[0009] 2) After the reaction is completed, the mixture is cooled and filtered, and the obtained filtrate is transferred to a dialysis bag. The dialyzed solution is freeze-dried to obtain magnesium-doped carbon dots.

[0010] The magnesium-doped carbon dots (Mg@CDs) provided by this invention incorporate both magnesium and nitrogen atoms into the carbon dots. This dual doping can further regulate the electronic structure and chemical reactivity of the carbon dots. The introduction of magnesium atoms can significantly alter the chemical properties of the surface or interior of the carbon dots, such as enhancing their photocatalytic activity. Fluorescence spectroscopy shows that the magnesium-doped carbon dots provided by this invention have an optimal excitation wavelength of 420 nm and an optimal emission wavelength of 650 nm. Under excitation with a handheld ultraviolet lamp, red light can be observed, significantly different from the blue light emitted by the magnesium-nitrogen-doped carbon dots disclosed in Chinese patent application CN 113201330 A.

[0011] Since magnesium hydroxide is difficult to dissolve, the present application first dissolves magnesium hydroxide alone in formamide and ultrasonically disperses it to obtain a reaction solution B, which is then reacted with the reaction solution A obtained by dissolving and dispersing tobacco powder and reduced glutathione. This allows the materials to be more fully dispersed and the quality of the magnesium-doped carbon dots obtained by the reaction is better.

[0012] Preferably, the reaction conditions include: pressure 0.2-3 MPa, temperature 160-190° C., and time 7-9 h. More preferably, the reaction conditions include: pressure 0.5-1.5 MPa, temperature 170-185° C., and time 7.5-8.5 h.

[0013] Preferably, the molecular weight cut-off of the dialysis bag is 3300-3700 Dalton. More preferably, the molecular weight cut-off of the dialysis bag is 3400-3600 Dalton.

[0014] Preferably, the dialysis time is 7-10 days. More preferably, the dialysis time is 7.5-9 days.

[0015] Preferably, the tobacco powder is obtained by removing stems, cutting, crushing and then sieving tobacco leaf samples.

[0016] In another aspect, the present invention provides the use of the magnesium-doped carbon dots in tobacco cultivation.

[0017] Preferably, the tobacco or tobacco seeds are in an abiotic stress environment, and the abiotic stress environment includes at least one of weak light, drought, and low temperature.

[0018] The present invention also provides a method for promoting tobacco seed germination, comprising the steps of preparing an aqueous solution of the magnesium-doped carbon dots at a concentration of 30-70 μg / mL and applying the solution to the tobacco seeds. Appropriate light exposure is required during germination, for example, 12 hours of light and 12 hours of darkness per day.

[0019] Preferably, the tobacco seeds are MS Yunyan 87 naked seeds.

[0020] Preferably, the concentration of the magnesium-doped carbon dots aqueous solution is 60 μg / mL.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] (1) The present invention provides a novel magnesium-doped carbon dot (Mg@CDs). Fluorescence spectroscopy shows that the optimal excitation wavelength of the magnesium-doped carbon dots provided by the present invention is 420 nm, and the optimal emission wavelength is 650 nm. These magnesium-doped carbon dots can increase the wavelength absorption range of tobacco seeds or tobacco plants for photosynthesis. Under the excitation of a handheld ultraviolet lamp, red light emission can be observed, which is significantly different from the blue light emission of magnesium-nitrogen-doped carbon dots disclosed in Chinese patent application CN 113201330A.

[0023] (2) The present invention provides novel magnesium-doped carbon dots for use in tobacco cultivation. Experimental studies have demonstrated that the magnesium-doped carbon dots provided by the present invention can effectively mitigate the effects of abiotic stress environments such as low light and drought, promoting tobacco seed germination with superior results compared to undoped magnesium carbon dots, thereby improving seedling cultivation. Under relatively extreme conditions, such as a low temperature of 10°C, 25% PEG simulated drought, and a weak light of 1000 lux, the magnesium-doped carbon dots provided by the present invention can also promote tobacco seed germination, resulting in faster germination and a higher germination index.

[0024] (3) The present invention also provides a method for promoting tobacco seed germination, wherein the concentration of the magnesium-doped carbon dot aqueous solution is low, which is beneficial to reducing costs. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 X-ray photoelectron spectrum of magnesium-doped carbon dots provided by the present invention; wherein 1-a is an analysis diagram of the entire magnesium-doped carbon dots, and 1-b is an analysis diagram of the elemental composition such as C and N.

[0026] Figure 2 Figure 1 shows the fluorescence spectrum detection results of magnesium-doped carbon dots provided by the present invention.

[0027] Figure 3 Observation diagram of magnesium-doped carbon dots provided by the present invention under excitation of a handheld ultraviolet lamp.

[0028] Figure 4 Germination index results of treatments with different concentrations of Mg@CDs.

[0029] Figure 5 Germination index results of treatments with different concentrations of CDs.

[0030] Figure 6 Effects of different light intensities on the germination rate of naked seeds of MS Yunyan 87.

[0031] Figure 7 Observation diagram of germination under different light intensities of Mg@CDs treatment.

[0032] Figure 8 Effects of different drought stress treatments on the naked seed germination rate of MS Yunyan 87.

[0033] Figure 9 Effects of different temperature treatments on the germination rate of naked seeds of MS Yunyan 87.

[0034] Figure 10 Comparison of carbon dots accumulated in tobacco plant leaf tissues after spraying.

[0035] Figure 11 Observation photo of MS Yunyan 87 tobacco plants 5 weeks after spraying. DETAILED DESCRIPTION

[0036] The present invention is further described in detail below with reference to the accompanying drawings and embodiments.

[0037] In this invention, the experimental materials involved are commonly available commercial products or can be obtained through conventional techniques in the art. The detection methods used are based on common methods in the art or the kit instructions. Unless otherwise specified, the percentage content is understood to be based on common usage in the art.

[0038] MS Yunyan 87 naked variety: MS Yunyan 87 was bred by the Yunnan Tobacco Science Research Institute and the China Tobacco Breeding Research (South) Center using Yunyan No. 2 as the female parent and K326 as the male parent. It was approved by the National Variety Approval Committee in December 2000.

[0039] The germination rate is the percentage of germinated seeds compared to the total number of seeds tested at the end of a specified germination test. The germination rate approximates the seedling rate, which is the ratio of the number of germinated seeds to the total number of seeds tested. The germination index is an indicator of seed vigor. During the germination test, the number of germinated seeds is recorded daily and the germination index is calculated. A higher germination index indicates higher seed vigor. The germination index reflects the germination rate; a higher germination index indicates greater seed vigor and faster seedling emergence under natural conditions.

[0040] Germination testing was conducted in accordance with the "Tobacco Industry Standard of the People's Republic of China YC / T 20-1994 - Tobacco Seed Inspection Procedure." Germination was observed and recorded at 4:00 PM each day. The specific recordings for different abiotic stress factors were as follows:

[0041] The germination rate of tobacco seeds was counted on the 14th day of the weak light stress experiment; the germination rate of tobacco seeds was counted on the 20th day of the drought stress experiment; and the germination rate of tobacco seeds was counted on the 24th day of the low temperature stress experiment.

[0042] Germination rate = total number of seeds that can form radicles or plumules / number of test seeds × 100%.

[0043] Germination index = GI = ΣGt / Dt (Dt is the number of days from the start of culture, Gt is the normal number of germinations on each corresponding day).

[0044] Data processing: Excel was used for preliminary data analysis and analysis, and DPS 7.05 was used for analysis of variance and multiple comparisons. A significance level of α = 0.05 and an extremely significant level of α = 0.01 were selected for hypothesis testing. Origin Pro 2018 was used for charting.

[0045] Example 1 Preparation of magnesium-doped carbon dots

[0046] The preparation method of magnesium-doped carbon dots (Mg@CDs) includes the following steps:

[0047] 1) 0.4 g of tobacco powder and 1.2 g of reduced glutathione were mixed, dissolved in 35 mL of formamide, and ultrasonically dispersed for 30 minutes to obtain reaction solution A; 0.52 g of magnesium hydroxide was dissolved in 25 mL of formamide, and ultrasonically dispersed for 30 minutes to obtain reaction solution B, which was mixed with reaction solution A and then placed in an autoclave for reaction. The reaction conditions included: pressure of 0.6-1 MPa, temperature of 180°C, and time of 8 hours;

[0048] 2) After the reaction is completed, the mixture is cooled and filtered, and the resulting filtrate is transferred to a dialysis bag with a molecular weight cutoff of 3500 Dalton. The solution after 8 days of dialysis is treated in a vacuum freeze dryer to obtain magnesium-doped carbon dots.

[0049] Example 2 Characterization of Mg-doped carbon dots

[0050] The magnesium-doped carbon dots prepared in Example 1 were subjected to X-ray photoelectron spectroscopy (XPS) analysis. The overall analysis of the magnesium-doped carbon dots is shown in FIG. Figure 1 -a, the analysis diagram of the elements such as C and N is as follows Figure 1 -b. Figure 1 It can be seen that the Mg element is successfully doped into the carbon dot nanomaterials.

[0051] The magnesium-doped carbon dots prepared in Example 1 were tested by fluorescence spectroscopy. Figure 2 As shown in Figure 2, the optimal excitation wavelength of magnesium-doped carbon dots is 420 nm, and the optimal emission wavelength is 650 nm. In addition, red light emission can be observed under the excitation of a handheld UV lamp, as shown in Figure 2. Figure 3 shown.

[0052] Example 3 Investigation of the Effect of Magnesium-doped Carbon Dots on the Germination of Naked Seeds of MS Yunyan 87

[0053] The preparation method of undoped magnesium carbon dots (CDs) for comparison includes the following steps: 1) mixing 0.05 g of tobacco powder with 0.30 g of reduced glutathione, adding 10 mL of formamide, and ultrasonically dispersing for 20 minutes. The mixture is then reacted in a muffle furnace at 180°C for 8 hours. 2) After the reaction, the mixture is cooled and centrifuged at a speed of 3500 r / min. The supernatant is filtered through an organic filter membrane with a pore size of 0.22 μm. The resulting filtrate is transferred to a dialysis bag with a molecular weight cutoff of 3500 Dalton and dialyzed for 7 days until no fluorescent substance is detected in the solution outside the dialysis bag. The dialyzed carbon dot solution is then filtered through an aqueous filter membrane with a pore size of 0.22 μm. The filtrate is freeze-dried to obtain undoped magnesium carbon dots.

[0054] 1 Experimental design

[0055] An experimental group and a control group were designed. Each group selected the same conditions but different treatment combinations (magnesium-doped carbon dot concentration, different light intensity, different temperature, and different simulated drought levels). The experiment was repeated 4 times, and one culture dish was used for each repetition, with 100 seeds sown in each culture dish.

[0056] 2 Investigation of the concentration of Mg-doped carbon dots

[0057] Tobacco seeds of uniform color and size were evenly placed in Petri dishes lined with cotton wool and filter paper. The dishes were then divided into four groups and incubated in a light-incubator, resulting in three experimental groups and one control group (CK). The concentrations of Mg@CDs (A1-A3) were set at 30.0 μg / mL, 60.0 μg / mL, and 90.0 μg / mL, respectively. Each experimental group underwent treatments (B1-B3), with Mg@CDs, CDs, and MgSO4 added to the tobacco seeds in the Petri dishes, respectively. Each treatment was replicated four times. The treatment settings for different Mg@CD concentrations are shown in Table 1.

[0058] Table 1 Treatment settings of different concentrations of Mg-doped carbon dots

[0059]

[0060] Each group in the experiment was treated with 12 hours of light and 12 hours of darkness every day. The light time period was: the first period: 8:00-20:00, temperature 25°C; the second period (darkness): 20:00-8:00 the next day, temperature 25°C; the relative humidity was maintained at 65%. The filter paper was always kept moist during the culture process, and seed germination was based on the radicle breaking through half the length of the seed. Starting from the second day after sowing, the number of seeds germinated and the growth of the seedlings after germination in the experimental group and the control group were recorded at the same time every day. Continue to observe for a period of time and record the relevant conditions until the seeds reach the maximum germination rate and end the experiment. The germination index results of different concentrations of Mg@CDs and CDs treatments are as follows: Figure 4 、 Figure 5 In the figure, lowercase letters indicate significant differences among different treatments at the P < 0.05 level, and uppercase letters indicate significant differences among different treatments at the P < 0.01 level.

[0061] from Figure 4 It can be seen that under the Mg@CDs treatment, there is no significant difference between 30.0μg / mL, 60.0μg / mL and 90.0μg / mL, and the difference between 0.0μg / mL and other treatments reaches an extremely significant level. Low concentrations of Mg@CDs can promote tobacco seed germination. As the concentration increases, the promotion effect weakens, but too high a concentration has a certain inhibitory effect on seed germination. At a concentration of 60.0μg / mL, the germination index of MS Yunyan 87 is the largest, reaching 34.59±1.47. Therefore, a concentration of 60.0μg / mL Mg@CDs was selected for subsequent experiments. Figure 5 It can be seen that although low concentrations of CDs can promote tobacco seed germination, there is no significant difference between 30.0μg / mL, 60.0μg / mL and 90.0μg / mL CDs and 0.0μg / mL.

[0062] 3. Lighting Experiment Investigation

[0063] Tobacco seeds of uniform color and size were evenly placed in Petri dishes lined with cotton wool and filter paper. The Petri dishes were then divided into four groups (C1-C4) and placed in a light incubator. Light intensities for C1-C4 were set at 1000 Lux, 4000 Lux, 7000 Lux, and 10,000 Lux. Each experimental group underwent four treatments, D1-D4. Treatments D1-D4 included ultrapure water (control), 60.0 μg / mL MgSO4, 60.0 μg / mL Mg@CDs, and 60.0 μg / mL CDs, respectively. Each treatment was replicated four times. The treatment settings for the different light intensity experiments are shown in Table 2.

[0064] Table 2 Treatment settings for different illumination experiments

[0065]

[0066]

[0067] Each group in the experiment was treated with 12 hours of light and 12 hours of darkness every day. The light time period was: the first period: 8:00-20:00, temperature 25°C; the second period (darkness): 20:00-8:00 the next day, temperature 25°C; the relative humidity was maintained at 65%. The filter paper was always kept moist during the cultivation process, and seed germination was based on the radicle breaking through half the length of the seed. Starting from the second day after sowing, the number of seeds germinated and the growth of the seedlings after germination of the experimental group and the control were recorded at the same time every day. Continue to observe for a period of time and record the relevant conditions until the seeds reach the maximum germination rate and end the experiment. The effects of different light intensity treatments on the germination rate and germination index of naked seeds of MS Yunyan 87 are shown as follows: Figure 6 , as shown in Table 3. The observation diagram of germination of Mg@CDs treated with different light intensities is shown in Table 3. Figure 7 As shown, 1, 2, 3, and 4 in the figure represent 4 repeated serial numbers.

[0068] Table 3 Effects of different light intensity treatments on the germination index of naked seeds of MS Yunyan 87

[0069]

[0070]

[0071] Duncan test: The data in the table are mean ± standard error; different lowercase letters indicate significant differences (P < 0.05), and different uppercase letters indicate extremely significant differences (P < 0.01).

[0072] Depend on Figure 6As can be seen, the seeds all began to germinate on the second day, reached their maximum germination rate on the fourth day, and gradually stabilized. On the second day after sowing, the number of seeds that germinated gradually increased with increasing light intensity. Furthermore, Mg@CDs was more effective than CDs, MgSO₄, and ultrapure water, while CDs was more effective than MgSO₄ and ultrapure water. Table 3 shows that under light intensities of 10,000 Lux, 7,000 Lux, and 4,000 Lux, the Mg@CDs and CDs carbon dot treatments showed extremely significant differences from MgSO₄ and ultrapure water, respectively, and there was also an extremely significant difference between Mg@CDs and CDs; however, there was no significant difference between MgSO₄ and ultrapure water. Under light intensities of 1,000 Lux, there was no significant difference between the Mg@CDs and CDs carbon dot treatments, but there were extremely significant differences between the MgSO₄ and ultrapure water. Under the same light intensity of 1000-10000 Lux, the germination index of tobacco seeds treated with carbon dots was greater than 1 compared to the control, indicating that carbon dots can improve the germination index of seeds. At 4000-10000 Lux, the germination index of tobacco seeds treated with Mg@CDs was significantly better than that of the CDs treatment. However, the germination index of tobacco seeds treated with MgSO4 was less than 1 compared to that of the ultrapure water treatment, possibly because the excessive concentration of 60 μg / mL MgSO4 inhibited seed germination.

[0073] Experimental investigation on 4PEG simulated drought stress

[0074] Tobacco seeds of uniform color and size were evenly placed in Petri dishes lined with cotton wool and filter paper. The Petri dishes were then divided into five groups (A1–A5) and placed in a light incubator. Experiments were conducted under different drought conditions (A1–A5). A1–A5 experiment simulated drought conditions by adding 30 mL of ultrapure water (normal), 15% PEG (mild drought), 20% PEG (moderate drought), 25% PEG (severe drought), and 30% PEG (extremely severe drought) to the Petri dishes, respectively. Each experimental group underwent four treatments (B1–B4), respectively, by adding 60.0 μg / mL Mg@CDs, 60.0 μg / mL CDs, 60.0 μg / mL MgSO₄, and ultrapure water (control) to the filter paper in the Petri dishes. Each treatment was replicated four times. The treatment settings for the different drought stress experiments are shown in Table 4.

[0075] Table 4 Treatment settings for different drought stress experiments

[0076]

[0077] Each group of the experiment was treated with 12 hours of light and 12 hours of darkness every day. The light time period was: the first period: 8:00-20:00, temperature 25°C; the second period (darkness): 20:00-8:00 the next day, light intensity of 10,000 Lux, temperature 25°C; relative humidity was maintained at 65%. The filter paper was always kept moist during the cultivation process, and seed germination was based on the radicle breaking through half the length of the seed. Recording began from the germination of seeds after sowing, and the number of seeds germinated and the growth of seedlings after germination of the experimental group and the control group were recorded at the same time every day. Continue to observe for a period of time and record relevant conditions until the seeds reach the maximum germination rate and end the experiment. The effects of different drought stress treatments on the germination rate and germination index of naked seeds of MS Yunyan 87 are shown as follows. Figure 8 , as shown in Table 5. Since the seeds did not germinate until they became moldy after being treated with 30% PEG, Figure 8 Results without 30% PEG treatment.

[0078] Table 5 Effects of different treatment combinations on the germination index of MS Yunyan 87

[0079]

[0080] Duncan test: The data in the table are mean ± standard error; different lowercase letters indicate significant differences (P < 0.05), and different uppercase letters indicate extremely significant differences (P < 0.01).

[0081] Depend on Figure 8 As can be seen, the seed germination lag period prolonged with increasing drought stress. At 0%, 15%, 20%, and 25% PEG concentrations, at the seed rupture stage, Mg@CDs was more effective than CDs, ultrapure water, and MgSO₄, and CDs was more effective than ultrapure water and MgSO₄. Table 5 shows that at the same PEG concentration, the germination index of the Mg@CDs and CDs carbon dot treatments was greater than 1, significantly different from the MgSO₄ and ultrapure water treatments, respectively. Furthermore, there was a significant difference between Mg@CDs and CDs, indicating that carbon dot materials can enhance seed germination index, with Mg@CDs significantly outperforming CDs in enhancing seed germination index. At 15% PEG concentration, MgSO₄ treatment was significantly superior to ultrapure water treatment. At 20% and 25% PEG concentrations, there was no significant difference between MgSO₄ and ultrapure water treatments.

[0082] 5 Temperature stress experimental investigation

[0083] Tobacco seeds of uniform color and size were evenly placed in Petri dishes lined with cotton wool and filter paper. The dishes were then divided into five groups (A1–A5) and placed in a light incubator. The experiments were conducted at different temperatures (A1–A5), with A1–A5 being 25°C, 20°C, 15°C, 10°C, and 5°C, respectively. Four treatments (B1–B4) were applied to each temperature, with 60.0 μg / mL Mg@CDs, 60.0 μg / mL CDs, 60.0 μg / mL MgSO₄, and ultrapure water (control). Each treatment was replicated four times. The treatment settings for the different temperature experiments are shown in Table 6.

[0084] Table 6 Treatment settings for experiments at different temperatures

[0085]

[0086]

[0087] Each group was treated with 12 hours of light and 12 hours of darkness every day. The light time period was as follows: the first period: 8:00-20:00, temperature 25°C; the second period (darkness): 20:00-8:00 the next day, relative humidity maintained at 65%, and light intensity of 10,000 Lux. The filter paper was always kept moist during the cultivation process, and seed germination was based on the radicle breaking through half the length of the seed. Recording began from the time the seeds germinated after sowing, and the number of seeds germinated and the growth of the seedlings after germination of the experimental group and the control group were recorded at the same time every day. Continue to observe for a period of time and record the relevant conditions until the seeds reach the maximum germination rate and end the experiment. The results of the effects of different temperature treatments on the germination rate and germination index of naked seeds of MS Yunyan 87 are as follows: Figure 9 , as shown in Table 7. Since the seeds did not germinate until they became moldy after being treated at 5℃, Figure 9 The results of the 5°C treatment are not included.

[0088] Table 7 Effects of different temperature treatments on the germination index of naked seeds of MS Yunyan 87

[0089]

[0090]

[0091] Duncan test: The data in the table are mean ± standard error; different lowercase letters indicate significant differences (P < 0.05), and capital letters indicate extremely significant differences (P < 0.01).

[0092] Depend on Figure 9As can be seen, seed germination times vary under different temperature conditions, and the lag phase of seed germination increases with decreasing temperature. At temperatures between 10 and 25°C, when the seeds break through the seed coat, Mg@CDs is more effective than CDs, ultrapure water, and MgSO4; CDs is more effective than ultrapure water. Table 7 shows that at the same temperature, the germination index of both Mg@CDs and CDs carbon dot treatments is greater than 1 compared to the ultrapure water treatment, indicating that carbon dots can enhance the seed germination index. The difference between Mg@CDs and CDs is highly significant at temperatures of 20 and 25°C. However, the germination index of tobacco seeds treated with MgSO4 is less than 1 compared to the ultrapure water treatment, likely due to the excessive concentration of 60 μg / mL MgSO4, which inhibits seed germination.

[0093] Example 4 Investigation of the Effect of Magnesium-doped Carbon Dots on the Growth of MS Yunyan 87 Tobacco Strain

[0094] This example further investigates the effect of spraying magnesium-doped carbon dot aqueous solution on the growth of MS Yunyan 87 tobacco plants. The carbon dots accumulated in the leaf tissues of tobacco plants after spraying are shown in the figure below. Figure 10 CK refers to the spraying of ultrapure water. Figure 11 As shown. Figure 10 and Figure 11 It can be seen that spraying magnesium-doped carbon dots aqueous solution can promote the accumulation of nutrients and the growth of tobacco plants.

[0095] The above is a further detailed description of the present invention in conjunction with specific preferred embodiments, and the specific implementation of the present invention should not be considered to be limited to these descriptions. For those skilled in the art of the present invention, without departing from the concept of the present invention, several simple deductions or substitutions can be made, which should be considered to fall within the scope of protection of the present invention.

Claims

1. A magnesium-doped carbon dot, characterized in that: The preparation method comprises the following steps: 1) Tobacco powder and reduced glutathione are mixed at a mass ratio of 1:(1.5-4), dissolved in formamide, and ultrasonically dispersed to obtain reaction solution A; magnesium hydroxide is dissolved in formamide at a mass ratio of 1.2-1.4 times that of the tobacco powder, and ultrasonically dispersed to obtain reaction solution B, which is mixed with reaction solution A and then placed in a high-pressure reactor for reaction; 2) After the reaction is completed, the mixture is cooled and filtered, and the obtained filtrate is transferred to a dialysis bag. The dialyzed solution is freeze-dried to obtain magnesium-doped carbon dots.

2. The magnesium-doped carbon dots according to claim 1, wherein: The reaction conditions include: pressure 0.2-3 MPa, temperature 160-190° C., and time 7-9 h.

3. The magnesium-doped carbon dots according to claim 1 or 2, wherein: The molecular weight cut-off of the dialysis bag is 3300-3700 Dalton.

4. The magnesium-doped carbon dots according to claim 1 or 2, characterized in that: The dialysis time is 7-10 days.

5. The magnesium-doped carbon dots according to claim 1 or 2, characterized in that: The tobacco powder is obtained by removing stems, cutting into shreds, crushing and then sieving tobacco leaf samples.

6. Use of the magnesium-doped carbon dots according to any one of claims 1 to 5 in tobacco cultivation.

7. The use of magnesium-doped carbon dots in tobacco planting according to claim 6, characterized in that: The tobacco or tobacco seeds are in an abiotic stress environment, and the abiotic stress environment includes at least one of weak light, drought, and low temperature.

8. A method for promoting tobacco seed germination, characterized in that: The method comprises the following steps: preparing the magnesium-doped carbon dots according to any one of claims 1 to 5 into an aqueous solution and applying the aqueous solution to tobacco seeds, wherein the concentration of the magnesium-doped carbon dots aqueous solution is 30-70 μg / mL.

9. The method for promoting tobacco seed germination according to claim 8, characterized in that: The tobacco seeds are naked varieties of MS Yunyan 87.

10. The method for promoting tobacco seed germination according to claim 8, characterized in that: The concentration of the magnesium-doped carbon dot aqueous solution is 60 μg / mL.

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