Nutrient solution for promoting potato seedling strengthening and enhancing stress resistance, culture method and application of nutrient solution

Abstract

The invention discloses a nutrient solution for promoting potato seedling strengthening and enhancing stress resistance, a culture method and application of the nutrient solution. Through a nutrient solution culture contrast test, the improved 1 / 2 Hoagland nutrient solution is found, short-term robustness of potato seedlings can be promoted by increasing the dosage of phosphorus and zinc elements, growth of tissue culture seedlings can be better promoted and the stress resistance of the tissue culture seedlings can be enhanced by adding seaweed oligosaccharide, only ionic liquid nutrient solution culture is needed, the method is suitable for water culture and matrix culture, the method is simple and rapid, and the cost is low. The strengthening promoting effect is good.

Description

Technical Field

[0001] The invention relates to the field of biotechnology, and in particular to a nutrient solution for promoting potato seedling growth and enhancing stress resistance, a culture method and application thereof. Background Art

[0002] Potato is the third largest food crop in the world and the fourth largest in my country. It is not only an important staple food crop, but also the most basic vegetable crop, an excellent feed crop, and an industrial raw material crop with huge value-added potential. Since potatoes are vegetatively propagated crops, they are mainly planted in tubers. Planting tubers for multiple generations can easily lead to the accumulation of viruses, viroids, bacteria, fungi and other pathogens in the tubers, which makes it impossible for potato plants to fully exert the production characteristics of the variety during the production process, resulting in a decrease in potato yield and quality. In order to overcome the above problems and prevent seed potatoes from "degenerating" with diseases, virus-free technology is currently used to produce virus-free seed potatoes for potato production, increase potato yield and improve quality. However, potato virus-free and factory-based production of virus-free seed potatoes cannot be separated from the cultivation of tissue culture seedlings.

[0003] At present, there are two main ways to propagate potato virus-free test tube seedlings. One is to propagate potato virus-free test tube seedlings directly in a matrix such as vermiculite, perlite, sand, sawdust, and base soil. This method takes a long time to propagate, has a low survival rate, is easily infected with pathogens, and results in a high production cost of the original seed. The other is to propagate potato virus-free test tube seedlings in a culture medium based on MS. This method has a relatively fast propagation speed and a relatively high survival rate, but has high production environment requirements and a high culture cost, which also results in a high production cost of the original seed. Therefore, it is urgent to develop a culture method and culture medium suitable for potato tissue culture seedling cultivation. Summary of the invention

[0004] The purpose of the present invention is to overcome the shortcomings and deficiencies of the prior art and provide a nutrient solution for promoting the growth of potato seedlings and enhancing stress resistance.

[0005] Another object of the present invention is to provide a cultivation method for promoting potato seedling growth and enhancing stress resistance.

[0006] Another object of the present invention is to provide an application of the above nutrient solution.

[0007] The purpose of the present invention is achieved through the following technical solutions:

[0008] A nutrient solution for promoting potato seedling growth and enhancing stress resistance, the components of which include: 4.0-6.0 mM KNO3, 2.0-3.0 mM Ca(NO3)2, 2.0-3.0 mM NH4Cl, 0.5-1.5 mM MgSO4, 0.5-1.5 mM KH2PO4, 15.0-25.0 μM H3BO3, 4.0-5.0 μM MnCl2, 1.0-1.4 μM ZnSO4, 0.1-0.2 μM CuSO4, 0.5-1.5 mM K2SO4, 80-100 μM FeSO4, 0.1-0.2 μM (NH4)6Mo7O 24 , 0.05~0.10mM Fe-Na-EDTA.

[0009] The components of the nutrient solution also include 0.033-0.400 g / L of seaweed oligosaccharide.

[0010] Preferably, the concentration of ZnSO4 in the components of the nutrient solution is 1.2 μM.

[0011] Preferably, the concentration of KH2PO4 in the components of the nutrient solution is 1.0 mM.

[0012] The nutrient solution is used in the cultivation of potato tissue culture seedlings, and can promote the rapid growth of the potato tissue culture seedlings and improve their stress resistance.

[0013] Application of the above nutrient solution in potato cultivation.

[0014] The potatoes are potato tissue culture seedlings.

[0015] Application of the above nutrient solution in enhancing potato stress resistance.

[0016] A method for cultivating strong potato tissue culture seedlings comprises the following steps:

[0017] Transplant the potato tissue culture seedlings into the growth medium and add nutrient solution for cultivation.

[0018] The potato tissue culture seedlings are tissue culture seedlings obtained by virus-free culture of potato tissues.

[0019] The growth medium is a nutrient solution or a peat soil matrix.

[0020] When the growth medium is peat soil matrix, the amount of nutrient solution added is twice a week, 100 mL each time.

[0021] The culture time is 20 to 60 days.

[0022] The culture temperature is 20-22°C.

[0023] The light intensity of the culture is 5 to 15 Klux.

[0024] The lighting time of the culture is 12 to 16 hours per day.

[0025] The culture solution is 1 / 2 Hoagland nutrient solution; preferably, it is 1 / 2 Hoagland nutrient solution added with seaweed oligosaccharides.

[0026] Compared with the prior art, the present invention has the following advantages and effects:

[0027] The potato tissue culture seedlings cannot grow up to the requirements of being transplanted to potted plants or fields due to the nutrient content of the growth medium and the space of the tissue culture bottle. In order to promote the potato tissue culture seedlings to grow rapidly and strong in a short period of time, the usual method is to change the culture medium composition, but due to the limitation of the tissue culture device and the difference in the environment, it is still impossible to expand the strong potato seedlings. Recently, it has been found that the growth of potato seedlings can be promoted by adding nutrient solution to vermiculite, but the nutrient solution composition of this method is complex, the planting process is cumbersome, and it is not ideal for promoting the growth of potato tissue culture seedlings. For this reason, the present invention has found a modified 1 / 2 Hoagland nutrient solution through a nutrient solution culture comparison test, and increasing the amount of phosphorus and zinc elements can promote the short-term strong growth of potato seedlings. Adding seaweed oligosaccharides can better promote the growth of tissue culture seedlings and enhance their stress resistance. Moreover, only ionic liquid nutrient solution needs to be used for culture, which is suitable for hydroponics and matrix culture. The method is simple and fast, and the effect of promoting strong growth is good. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 It is a potato plant in the hydroponic environment and cultivation test in the embodiment.

[0029] Figure 2 This is a graph showing the plant yield results of the culture experiment with different culture solutions in Example 1.

[0030] Figure 3 This is a graph showing the dry weight results of plants cultured in different culture solutions in Example 1.

[0031] Figure 4 This is a graph showing the results of the culture experiment in Example 3 in which the phosphorus concentration was adjusted alone.

[0032] Figure 5 This is a graph showing the results of the culture experiment in Example 3 in which the zinc concentration was adjusted alone.

[0033] Figure 6 This is a graph showing the results of a culture test in Example 4 in which different concentrations of seaweed oligosaccharides were added to the improved culture solution.

[0034] Figure 7 1 is a graph showing the results of the culture experiment at different light intensities in Example 5.

[0035] Figure 8This is a graph showing the results of detecting the expression levels of related enzymes in plants cultured in different improved culture solutions after low temperature treatment in Example 6.

[0036] Fig. 9 This is a graph showing the results of detecting the expression levels of related enzymes in plants cultured in different improved culture solutions after high temperature treatment in Example 6.

[0037] Fig.10 It is a graph showing the results of detecting the expression levels of related genes in plants cultured in different improved culture solutions after low temperature treatment in Example 6; wherein positive values ​​represent the results of each treatment compared with treatment T9, and negative values ​​represent the expression results of treatment T9 compared with each other treatment.

[0038] Fig.11 This is a graph showing the results of detecting the expression levels of related genes in plants cultured in different improved culture solutions after high temperature treatment in Example 6; wherein positive values ​​represent the results of each treatment compared with treatment T9, and negative values ​​represent the expression results of treatment T9 compared with each other treatment. DETAILED DESCRIPTION

[0039] The present invention is further described in detail below in conjunction with embodiments and drawings, but the embodiments of the present invention are not limited thereto.

[0040] If no specific experimental conditions are specified in the following embodiments, conventional experimental conditions or experimental conditions recommended by the reagent company are generally followed. The materials and reagents used, unless otherwise specified, are all reagents and materials obtained from commercial sources.

[0041] Example 1 Screening of basal culture fluid

[0042] 1.1 Cultivation of potato tissue culture seedlings

[0043] Prepare the culture medium with 4.43g / L MS powder, 30g / L sucrose, 15g / L agar, pH 5.8-6.0, sterilize at 121℃ for 20min, and dispense into tissue culture bottles. Move the single-node potato stem segment with axillary buds into the tissue culture bottle containing the culture medium, place it in a light culture at 20-22℃ and 8-10Klux for 14 hours, and observe the tissue culture seedlings after 4-6 weeks. When the height is about 2 / 3 of the height of the tissue culture bottle, it can be used for transplanting. 1.2 Basic culture medium seedling culture experiment

[0044] The potato tissue culture seedlings obtained in 1.1 were transplanted into an aeroponic box for hydroponic culture. The potato plants were cultured using four different nutrient solutions, including 1 / 2 Hoagland, MS, GSM and PD. The culture temperature was 22°C, the light intensity was 8-10Klux, and the lighting time was 16 hours. After 60 days of culture, the whole plant was collected, the mass of the tubers was measured to calculate the yield, and the plants were weighed to calculate the dry weight.

[0045] Treatment 1: 1 / 2 Hoagland medium, which consists of 5.0 mM KNO3, 2.5 mM Ca(NO3)2, 2.5 mM NH4Cl, 1.0 mM MgSO4, 0.5 mM KH2PO4, 20 μM H3BO3, 4.5 μM MnCl2, 1.0 μM ZnSO4, 0.16 μM CuSO4, 1.0 mM K2SO4, 90 μM FeSO4, 0.16 μM (NH4)6Mo7O 24 and 0.08 mM Fe-Na-EDTA;

[0046] Treatment 2: MS culture medium, whose composition is 37.5mM KNO3, 4.0mM Ca(NO3)2, 20.6mM NH4Cl, 1.7mMMgSO4, 1.25mM KH2PO4, 100μM H3BO3, 100μM MnCl2, 25μM ZnSO4, 0.25μM CuSO4, 0.01mM K2SO4, 1.0μM (NH4)6Mo7O 24 and 0.08 mM Fe-Na-EDTA;

[0047] Treatment 3: GSM medium, whose composition is 28.5mM KNO3, 0.8mM Ca(NO3)2, 2.5mM NH4Cl, 1.2mMMgSO4, 1.25mM KH2PO4, 84μM H3BO3, 10μM MnCl2, 3.0μM ZnSO4, 2.6μM CuSO4, 0.07mM K2SO4, 90μM FeSO4, 2.8μM (NH4)6Mo7O 24 and 0.08 mM Fe-Na-EDTA;

[0048] Treatment 4: PD culture medium, whose composition is 15mM KNO3, 2.5mM Ca(NO3)2, 3.0mM NH4Cl, 1.5mMMgSO4, 1.0mM KH2PO4, 60μM H3BO3, 9.0μM MnCl2, 2.0μM ZnSO4, 0.8μM CuSO4, 0.2mM K2SO4, 45μM FeSO4, 1.0μM (NH4)6Mo7O 24 and 0.08 mM Fe-Na-EDTA;

[0049] The experimental results are as follows Figure 2 and Figure 3As shown, treatment 1 had the highest potato yield, with an average of 278g per plant, followed by treatment 2 with 216g per plant, and treatment 4 had the lowest yield, with a single plant weight of 16g. Treatment 1 had the highest dry weight of potato plants, with an average dry weight of 34.2g per plant, followed by treatment 4 with a single plant dry weight of 26.7g, and treatment 2 had the lowest dry weight, with a single plant dry weight of 18.6g. This proved that 1 / 2 Hoagland culture solution had the best seedling strengthening effect, and 1 / 2 Hoagland culture solution was used as the basic culture solution for subsequent experiments.

[0050] Example 2 Screening of Growth Medium

[0051] 2.1 Experiment on seedling cultivation in different growth media

[0052] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, cultured in 1 / 2 Hoagland culture solution, transplanted to different growth media for treatment and culture, and collected and tested after 30 days of natural light culture.

[0053] Treatment 1: agar + 1 / 2 Hoagland medium, tissue culture bottle growth;

[0054] Treatment 2: nutrient-free peat soil substrate + 1 / 2 Hoagland nutrient solution, supplement nutrient solution twice a week, 100 mL each time;

[0055] Treatment 3: Cotton wool + 1 / 2 Hoagland nutrient solution, use cotton wool to fix the tissue culture seedlings in the hydroponic bread box, and culture the tissue culture seedlings in liquid form.

[0056] The experimental results are shown in Table 1. After the tissue culture seedlings under the three treatments grew for 30 days under the same growth environment, treatment 3 had significant advantages in root length, root number, plant height, plant internode number, leaf number and plant dry weight. Compared with treatment 1, treatment 3 significantly increased root length, root number, plant height, plant internode number, leaf number and plant dry weight by 35.2%, 57.1%, 47.2%, 29.2%, 61.4% and 74.4%, respectively. This shows that treatment 3 is more suitable for the growth of potato tissue culture seedlings, and the robustness of the seedlings is the best. However, treatment 3 cannot simulate the real growth environment in the field. The buffering effect of the soil matrix on the temperature is different from the culture solution environment for the growth of potato seedlings. In addition, hydroponics requires complex equipment such as aerosol boxes, which is complex and costly to operate, and is not suitable for mass production and promotion. In order to more comprehensively verify the experimental results, subsequent experiments used hydroponics and soil matrix as growth media to verify their seedling-strengthening effects.

[0057] Table 1 Results of seedling cultivation experiments in different growth media

[0058] deal with Root length / cm Number of roots Plant height / cm Number of internodes Robustness Number of blades Plant dry weight / g Process 1 16.5 4.2 10.6 4.8 Poor 12.7 1.25 Process 2 18.4 5.9 12.4 5.4 Strong 16.9 1.59 Process 3 22.3 6.6 15.6 6.2 robust 20.5 2.18

[0059] 2.2 Effects of different nutrient solution formulas on potato growth in matrix tissue culture seedling cultivation

[0060] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, transplanted into pots with peat soil matrix as the growth medium, and cultured with different nutrient solutions. The nutrient solution was supplemented twice a week, 100 mL each time, and an equal amount of water was supplemented according to growth needs. Each treatment was managed the same, exposed to natural light, and collected and tested after 40 days of cultivation.

[0061] Treatment 1: substrate + commercially available conventional amount of compound fertilizer (15-15-15);

[0062] Treatment 2: substrate + 1 / 2 Hoagland nutrient solution;

[0063] Treatment 3: substrate + MS nutrient solution;

[0064] Treatment 4: substrate + GSM nutrient solution;

[0065] Treatment 5: substrate + PD nutrient solution.

[0066] The experimental results are shown in Table 2. Comparing the various treatments, it was found that treatment 2 had the longest root length, plant height, number of leaves and plant dry weight, and treatment 2 had the best robustness, indicating that the nutrient solution scheme of treatment 2 was better than the traditional compound fertilizer scheme, and among the four nutrient solution schemes, the nutrient solution scheme of treatment 2 was the best.

[0067] Table 2 Effects of different nutrient solution formulas on potato growth in matrix tissue culture seedling cultivation

[0068]

[0069] Example 3 Effect of different phosphorus / zinc dosages on the growth of potato tissue culture seedlings

[0070] 3.1 Experiment on adjusting phosphorus / zinc dosage to strengthen seedlings

[0071] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, and transplanted into an aeroponic chamber for culture in 1 / 2 Hoagland culture solution containing different concentrations of phosphorus / zinc under natural light. After 30 days of culture, the seedlings were collected and tested.

[0072] Phosphorus treatment: 1 / 2 Hoagland nutrient solution only changed the phosphorus concentration, and the concentrations of other elements remained unchanged. The phosphorus concentrations were 0.5, 1.0, and 1.5 mM respectively;

[0073] Zinc treatment: 1 / 2 Hoagland nutrient solution only changed the zinc concentration, while the concentrations of other elements remained unchanged. The zinc concentrations were 1.0, 1.2, and 1.4 μM, respectively.

[0074] The experimental results are as follows Figure 4 and Figure 5 As shown in the figure, different phosphorus concentration treatments significantly affected the plant height, branch number, fresh weight and dry weight of potato tissue culture seedlings. Compared with P0.5, the plant height and plant fresh weight of P1.0 and P1.5 increased significantly, but there was no significant difference between P1.0 and P1.5. The dry weight of the plant was measured and it was found that the dry weight of the potato in the P1.0 treatment increased significantly, reaching a maximum of 1.74g, followed by the treatment P1.5, indicating that the phosphorus concentration P1.0 is more suitable for the growth of potato tissue culture seedlings. Similarly, compared with Zn1.0, Zn1.2 and Zn1.4 significantly increased the fresh weight and dry weight of potato plants, respectively, but there was no significant difference in plant height and root length, and there was no significant difference between Zn1.2 and Zn1.4 in each measured index, indicating that the zinc concentration Zn1.2 is more suitable for the cultivation of potato tissue culture seedlings.

[0075] 3.2 Simultaneous adjustment of phosphorus and zinc dosage to strengthen seedling cultivation experiment

[0076] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, transplanted into a hydroponic box, cultured in 1 / 2 Hoagland culture solution containing different concentrations of phosphorus / zinc, exposed to natural light, and collected and tested after 25 days of culture.

[0077] Treatment 1: 1 / 2 Hoagland nutrient solution;

[0078] Treatment 2: zinc + 1 / 2 Hoagland nutrient solution was added, and the zinc treatment was changed from 1.0 μM ZnSO4 in treatment 1 to 1.2 μM ZnSO4;

[0079] Treatment 3: phosphorus + 1 / 2 Hoagland nutrient solution was added, and the phosphorus treatment was changed from 0.5mM KH2PO4 in treatment 1 to 1.0mM KH2PO4;

[0080] Treatment 4: Zinc and phosphorus + 1 / 2 Hoagland nutrient solution were added at the same time. The zinc treatment was changed from 1.0μM ZnSO4 in treatment 1 to 1.2μM ZnSO4; the phosphorus treatment was changed from 0.5mM KH2PO4 in treatment 1 to 1.0mM KH2PO4.

[0081] The experimental results are shown in Table 3. After 25 days, the root length, lateral root number, plant height, stem thickness, leaf number and plant dry weight of the improved nutrient solution treatment 2, treatment 3 and treatment 4 were significantly increased under the same growth environment conditions. The observation of stem thickness, leaf number and dry weight showed that compared with treatment 1, treatment 2, treatment 3 and treatment 4 increased by 3.3%, 4.9%, 17.9%, 2.8%, 6.2%, 13.5%, 5.2%, 15.6% and 30.7% in stem thickness, leaf number and dry weight, respectively. It can be found from the data that increasing zinc or phosphorus concentration in treatment 2 and treatment 3 can promote potato stem thickness, increase leaf number and increase plant biomass. After treatment 4 increased zinc and phosphorus concentrations at the same time, the effect of promoting the growth of potato tissue culture seedlings was more obvious, indicating that zinc-phosphorus interaction can promote the growth of tissue culture seedlings, so the following experiment uses the improved phosphorus and zinc concentrations for further research.

[0082] Table 3 Results of the experiment on simultaneously adjusting the phosphorus / zinc dosage to strengthen the seedling culture

[0083]

[0084] 3.3 Interaction between phosphorus and zinc in substrate culture promotes the growth of potato seedlings

[0085] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, transplanted into a matrix growth medium, and cultured with different nutrient solutions under natural light. After 30 days of culture, the seedlings were collected and tested.

[0086] Treatment 1: substrate + clean water;

[0087] Treatment 2: substrate + 1 / 2 Hoagland nutrient solution;

[0088] Treatment 3: substrate + zinc + 1 / 2 Hoagland nutrient solution, the zinc treatment was changed from 1.0 μM ZnSO4 in treatment 1 to 1.2 μM ZnSO4;

[0089] Treatment 4: substrate + phosphorus + 1 / 2 Hoagland nutrient solution, the phosphorus treatment was changed from 0.5mM KH2PO4 in treatment 1 to 1.0mM KH2PO4 (same as treatment 3 in 3.2);

[0090] Treatment 5: substrate + zinc + phosphorus + 1 / 2 Hoagland nutrient solution, the zinc treatment was changed from 1.0 μM ZnSO4 in treatment 1 to 1.2 μM ZnSO4; the phosphorus treatment was changed from 0.5 mM KH2PO4 in treatment 1 to 1.0 mM KH2PO4 (same as treatment 4 in 3.2).

[0091] The experimental results are shown in Table 4. Compared with treatment 1, the root length, lateral root number, plant height, stem thickness, leaf number and plant dry weight of nutrient solution treatments 2, 3, 4 and 5 increased significantly after 30 days under the same growth environment. Observation of stem thickness, leaf number and dry weight showed that compared with treatment 2, treatments 3, 4 and 5 increased by 6.0%, 0.6%, 20.6%, 10.0%, 13.3%, 42.2%, 28.0%, 15.2% and 68.6% in stem thickness, leaf number and dry weight, respectively. Analysis of the data showed that increasing zinc or phosphorus concentration in treatments 3 and 4 could promote potato stem thickness, increase leaf number and increase plant biomass. After increasing zinc and phosphorus concentrations at the same time in treatment 5, the effect of promoting the growth of potato tissue culture seedlings was more obvious, indicating that the interaction between zinc and phosphorus can promote the growth of tissue culture seedlings, which is consistent with the results of hydroponic research.

[0092] Table 4 Substrate culture + changing the amount of phosphorus and zinc to promote the growth of potato tissue culture seedlings

[0093]

[0094] Example 4 Effect of adding different dilution multiples of seaweed oligosaccharides to improve nutrient solution on the growth of potato seedlings

[0095] 4.1 Adding different dilution multiples of seaweed oligosaccharides under hydroponic conditions

[0096] Potato tissue culture seedlings were cultured according to the method of 1.1 in Example 1, transplanted into a hydroponic box, cultured with different nutrient solutions, and added with seaweed oligosaccharides in the experimental group (the product name is brown algae oligosaccharide, the specification is 99%, purchased from Xi'an Yatu Biotechnology Co., Ltd., the seaweed oligosaccharide powder was configured into a 20 g / L stock solution, and added to the nutrient solution at a final concentration of 50 to 600 times dilution), exposed to natural light, and collected and tested after culturing for 22 days.

[0097] Treatment 1: deionized water culture and normal hydroponic culture;

[0098] Treatment 2: nutrient solution 1 / 2 Hoagland-i (same as treatment 4 in 3.2 of Example 3) was hydroponically cultured normally;

[0099] Treatment 3: nutrient solution 1 / 2 Hoagland-i + 0.400 g / L seaweed oligosaccharide;

[0100] Treatment 4: nutrient solution 1 / 2 Hoagland-i + 0.200 g / L seaweed oligosaccharide;

[0101] Treatment 5: nutrient solution 1 / 2 Hoagland-i + 0.100 g / L seaweed oligosaccharide;

[0102] Treatment 6: nutrient solution 1 / 2 Hoagland-i + 0.067 g / L seaweed oligosaccharide;

[0103] Treatment 7: nutrient solution 1 / 2 Hoagland-i + 0.050 g / L seaweed oligosaccharide;

[0104] Treatment 8: nutrient solution 1 / 2 Hoagland-i + 0.040 g / L seaweed oligosaccharide;

[0105] Treatment 9: nutrient solution 1 / 2 Hoagland-i + 0.033 g / L seaweed oligosaccharide;

[0106] The experimental results are shown in Table 5. Treatment 4 had the best effect. Compared with treatment 2, the plant height, number of leaves, stem diameter and dry weight increased by 27.27%, 41.75%, 21.11% and 93.65%, respectively, indicating that treatment 4 with the addition of 0.200 g / L seaweed oligosaccharides was more suitable for the growth of potato seedlings.

[0107] Table 5 Effects of different dilution multiples of seaweed oligosaccharides and improved nutrient solution on the growth of potato seedlings

[0108]

[0109] Note: The growth rate is compared with treatment 2

[0110] 4.2 Effects of different dilution multiples of seaweed oligosaccharide treatment and improved nutrient solution on the growth of potato seedlings in substrate cultivation

[0111] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, transplanted into an aeroponic box, using the matrix as the growth medium, and poured with different nutrient solutions for culture, exposed to natural light, and collected and tested after 26 days of culture.

[0112] Treatment 1: normal culture in hydroponics with substrate + deionized water;

[0113] Treatment 2: substrate + nutrient solution 1 / 2 Hoagland-i (modified to 1.2 μM ZnSO4 for zinc and 1.0 mM KH2PO4 for phosphorus after experiment) in normal hydroponic culture;

[0114] Treatment 3: substrate + nutrient solution 1 / 2 Hoagland-i + 0.400 g / L seaweed oligosaccharide;

[0115] Treatment 4: substrate + nutrient solution 1 / 2 Hoagland-i + 0.200 g / L seaweed oligosaccharide;

[0116] Treatment 5: substrate + nutrient solution 1 / 2 Hoagland-i + 0.100 g / L seaweed oligosaccharide;

[0117] Treatment 6: substrate + nutrient solution 1 / 2 Hoagland-i + 0.067 g / L seaweed oligosaccharide;

[0118] Treatment 7: substrate + nutrient solution 1 / 2 Hoagland-i + 0.050 g / L seaweed oligosaccharide;

[0119] Treatment 8: substrate + nutrient solution 1 / 2 Hoagland-i + 0.040 g / L seaweed oligosaccharide;

[0120] Treatment 9: substrate + nutrient solution 1 / 2 Hoagland-i + 0.033 g / L seaweed oligosaccharide;

[0121] The experimental results are as follows Figure 6 As shown, treatment 4 had the best effect. Compared with treatment 2, plant height, leaf number, stem diameter and dry weight increased by 20.0%, 28.2%, 24.3% and 98.65%, respectively. The differences in stem diameter and dry weight were significant, indicating that treatment 4 with the addition of 0.200 g / L seaweed oligosaccharides was also suitable for the growth of potato seedlings in substrate cultivation.

[0122] Example 5 Effect of different light intensities on the growth of potatoes cultured in nutrient solution supplemented with seaweed oligosaccharides

[0123] 5.1 Cultivation experiment under different light intensities under hydroponic conditions

[0124] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, transplanted under hydroponic conditions, and cultured using the culture solution of Treatment 4 in Example 4. Different light intensities were set and the seedlings were collected and tested after culturing for 18 days.

[0125] Light treatment L1: light intensity from the light source to the top of the plant growth is 5Klux;

[0126] Light treatment L2: light intensity from the light source to the top of the plant growth is 10Klux;

[0127] Light treatment L3: light intensity from the light source to the top of the plant growth is 15Klux;

[0128] The experimental results are as follows Figure 7 As shown in the figure, different light intensity treatments significantly affected the plant height, stem diameter and dry weight of potato tissue culture seedlings. Compared with L1, the plant height and stem diameter of L2 and L3 increased significantly, but there was no significant difference between L2 and L3 in terms of plant height. The dry weight of the plants was measured and it was found that the dry weight of potatoes in the L2 treatment increased significantly, and the fresh weight of the plants in the L2 treatment increased by 96.7% and 35.9% respectively compared with the treatments L1 and L3, indicating that the light intensity L2 is more suitable for the growth of potato tissue culture seedlings.

[0129] 5.2 Culture experiment under different light intensities under matrix culture conditions

[0130] Potato tissue culture seedlings were obtained by culturing according to the method of 1.1 in Example 1, transplanted into a matrix growth medium, and poured with the culture solution of Treatment 4 in Example 4. Different light intensities were set, and the seedlings were collected and tested after culturing for 23 days.

[0131] Light treatment L1: light intensity from the light source to the top of the plant growth is 5Klux;

[0132] Light treatment L2: light intensity from the light source to the top of the plant growth is 10Klux;

[0133] Light treatment L3: light intensity from the light source to the top of the plant growth is 15Klux;

[0134] The experimental results are shown in Table 6. Different light intensity treatments significantly affected the plant height, stem diameter and dry weight of potato tissue culture seedlings. Compared with L1, the plant height and stem diameter of L2 and L3 increased significantly, with plant height and stem diameter growth rates of 38.26%, 146.03% and 15.25%, 119.86%, respectively. The dry weight of the plants was measured and it was found that the dry weight of potatoes in the L2 treatment increased significantly, and the fresh weight of the plants in the L2 treatment increased by 103.2% and 13.4%, respectively, compared with the treatments L1 and L3, indicating that the light intensity L2 is more suitable for the growth of potato tissue culture seedlings in substrate cultivation.

[0135] Table 6 Effects of different light intensities on the improvement of nutritional culture of tissue culture seedlings by adding seaweed oligosaccharides

[0136]

[0137] Example 6 Comprehensive evaluation of phosphorus and zinc nutrition and seaweed oligosaccharides on high and low temperature resistance

[0138] According to the results of the previous experiments, it can be seen that adjusting the content of phosphorus / zinc and seaweed oligosaccharides in the nutrient solution has a significant effect on the growth of potato seedlings. In order to further study the effects of different culture solution combinations on potato seedlings, this experiment conducted experimental tests on the stress resistance of potato seedlings.

[0139] The potato tissue culture seedlings were cultured according to the method of 1.1 in Example 1, and transplanted into a light incubator. The light intensity of the light source to the top of the plant growth was 10 Klux. The matrix was used as the growth medium and different nutrient solutions were poured into the culture. After 28 days of culture, a control group, a low temperature treatment group and a high temperature treatment group were set for each treatment. After the treatment, the potato plant leaves were collected and stored at -80°C for detecting the leaf enzyme activity and the expression of purple acid phosphatase-related genes. Three replicates were set for each group.

[0140] Treatment T1: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.2+P1.0) + 0.200 g / L seaweed oligosaccharide;

[0141] Treatment T2: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.2+P0.5) + 0.200 g / L seaweed oligosaccharide;

[0142] Treatment T3: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.2+P1.5) + 0.200 g / L seaweed oligosaccharide;

[0143] Treatment T4: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.0+P0.5) + 0.200 g / L seaweed oligosaccharide;

[0144] Treatment T5: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.4+P0.5) + 0.200 g / L seaweed oligosaccharide;

[0145] Treatment T6: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.2+P1.0) + 0.400 g / L seaweed oligosaccharide;

[0146] Treatment T7: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.2+P1.0) + 0.050 g / L seaweed oligosaccharide;

[0147] Treatment T8: substrate + 0.200 g / L seaweed oligosaccharide;

[0148] Treatment T9: substrate + nutrient solution 1 / 2 Hoagland-i (Zn1.2+P1.0).

[0149] The Zn concentration unit of the above nutrient solution is μmol, the P concentration unit is mmol, and the other nutrient concentrations refer to the nutrient solution 1 / 2 Hoagland.

[0150] Low temperature treatment: The treatment temperature was -2°C. Leaf samples were collected 5 hours after treatment for enzyme activity analysis (Solabo kit) and gene expression level detection.

[0151] High temperature treatment: The treatment temperature was 38°C. After 5 hours of treatment, leaf samples were collected for enzyme activity analysis (Solabo kit) and gene expression detection.

[0152] Enzyme activity detection: The analysis method of enzyme activity POD, CAT, MDA, SOD and Pro was determined according to the operation method of Solebo kit ( Science & Technology Co. Beijing, China).

[0153] Analysis of gene expression: In order to more comprehensively evaluate the resistance changes of potatoes after seedling strengthening treatment with different culture media, the expression level of purple acid phosphatase (PAP), a related gene that plays an important role in the stress resistance process in the collected samples, was analyzed. The PAP gene is an important gene that responds to phosphorus starvation and activates the utilization of organic phosphorus sources inside and outside the plant, and is involved in regulating stress and hormone changes. According to the potato genome information website the Spud DB Potato Genomics Resources (DM v6.1, http: / / spuddb.uga.edu) (Rudic, J., Dragicevic, MB, Momcilovic, I., Simonovic, AD, Pantelic, D., 2022. In Silico Study of Superoxide Dismutase Gene Family in Potato and Effects of Elevated Temperature and Salicylic Acidon Gene Expression. Antioxidants 11(3), 488.), the key purple acid phosphatase (PAP)-related genes retrieved are Soltu.DM.01G049890, Soltu.DM.01G049900, Soltu.DM.03G019440, Soltu.DM.03G022790, Soltu.DM.04G003760 and Soltu.DM.04G036010. Primers were designed for the six PAP genes retrieved using the bioinformatics software Primer Premier 6.0, and the expression changes of genes were detected by real-time fluorescence quantitative PCR (qRT-PCR). RNA extraction and reverse transcription from plant leaves were performed according to the soybean RNA extraction method (Chengchen, Li, Shunhua, Gui, Tao, & Yang, et al. (2011). Identification of Soybean Purple Acid Phosphatase Genes and Their Expression Responses to Phosphorus Availability and Symbiosis. International Symposium on Integrative Plant Biology.).All samples were analyzed on a CFX96 real-time PCR detection system (Bio-Rad, 226 Hercules, USA). The potato housekeeping gene EF1A (F: 5'-227ATTGGAAACGGATATGCTCCA-3'; R: 5'-TCCTTACCTGAACGCCTGTCA-3') was used as an endogenous control for normalization. qRT-PCR used a 20 μL reaction mixture, including 2 μL cDNA template, 0.8 μL forward primer, 0.8 μL reverse primer, 10 μL 2×SYBR qPCR Master mix, and 6.4 μL RNase-free H2O. The thermal cycling conditions were as follows: initial denaturation at 95°C for 2 min, followed by denaturation at 95°C for 15 s, collection of fluorescence signals at 60°C for 30 s, and repeated 40 cycles. Each experimental treatment included 4 biological replicates, and the gene expression level was 2. -ΔCt Method calculation (Wang, Y., Li, Y., Zhou, F., Zhang, L., Gong, J., Cheng, C. et al., 2023. Genome-wide characterization, phylogenetic and expression analysis of Histone gene family in cucumber (Cucumis sativus L.). Int. J. Biol. Macromol. 230, 123401.).

[0154] The experimental results are shown in Table 7 and Figures 8 to 11 The data in Table 7 show that different nutrient solutions and seaweed oligosaccharide treatments affect the plant height, stem diameter and dry weight of potato tissue culture seedlings. In contrast, treatment 1 has the largest data of stem diameter, leaf area, biomass and root-to-crown ratio, which greatly promotes the growth of potato tissue culture seedlings.

[0155] Table 7 Effects of different nutrient solution concentrations and seaweed oligosaccharides on potato growth

[0156]

[0157] like Figure 8 As shown in the figure, the enzyme activities of different nutrient solutions and seaweed oligosaccharides changed differently after 5 hours of low temperature treatment. As can be seen from the figure, treatment 1 has higher POD, CAT, MDA, SOD and Pro enzyme activities, indicating that the treatment nutrient solution combination improves the low temperature resistance compared with other treatments.

[0158] like Fig. 9As shown in the figure, the enzyme activities of different nutrient solutions and seaweed oligosaccharides changed differently after 5 hours of high temperature treatment. As can be seen from the figure, treatment 1 has higher POD, CAT, MDA, SOD and Pro enzyme activities, indicating that the nutrient solution combination of treatment 1 has improved the high temperature resistance compared with other treatments.

[0159] Fig.10 qRT-PCR analysis of the expression of potato PAP-related genes showed that after 5 hours of treatment at -2℃, compared with the control treatment T9, under the same phosphorus and zinc nutrients, the addition of different concentrations of seaweed oligosaccharides to treatments T1, T6 and T7, the genes Soltu.DM.01G049890 and Soltu.DM.01G049900 showed the largest negative increase in gene expression in the leaves of T1 treatment, indicating that under the treatments of adding seaweed oligosaccharides to treatments T1, T6 and T7, the expression of PAP was significantly reduced compared with the control treatment T9 without adding seaweed oligosaccharides, and the reduction in treatment T1 was the largest, revealing that under the T1 treatment, potato plants obtained the least utilization of leaf organic phosphorus while meeting the growth and obtaining maximum biomass, while compared with treatment T1, the demand for organic phosphorus utilization in treatments T6 and T7 was increasing. Similarly, the expression of T1 treatment in Soltu.DM.03G019440 and Soltu.DM.03G022790 increased by the least fold, and the expression of T6 and T7 treatments was higher than that of T1 treatments, indicating that the expression of genes in organic phosphorus utilization in T1 treatment was lower. The expression of these four genes all showed that under chilling stress, the addition of seaweed oligosaccharides in T1 treatment increased the effectiveness of phosphorus nutrients and enhanced the resistance of potato plants to chilling stress.

[0160] Fig.11The qRT-PCR analysis of the expression of PAP-related genes in potato showed that after 5 hours of treatment at 38℃, compared with the control treatment T9, under the same phosphorus and zinc nutrients, the addition of different concentrations of seaweed oligosaccharides in treatments T1, T6 and T7, the gene Soltu.DM.01G049890, Soltu.DM.01G049900 and Soltu.DM.04G036010 in T1 treatment had the largest negative increase in expression times, indicating that seaweed oligosaccharides may be involved in regulating the phosphorus nutrition process of potatoes. The expression of Soltu.DM.01G049890 was significantly lower than that of the control treatment T9 without adding seaweed oligosaccharides, and the reduction in treatment T1 was the largest, revealing that under the T1 treatment, the potato plants obtained the least organic phosphorus utilization when meeting the growth and obtaining the maximum biomass, while the demand for organic phosphorus in treatments T6 and T7 was increasing compared with treatment T1. Similarly, Soltu.DM.01G049900 and Soltu.DM.04G036010 had the highest expression reduction in T1 treatment, and T6 and T7 had higher expression increase than T1 treatment, indicating that under the T1 nutrient combination, the expression of PAP genes for the acquisition and utilization of organic phosphorus in potato plant growth was low. Compared with T9, the expression of Soltu.DM.04G022790 in T1 treatment also increased the least. In short, the expression of these four genes consistently showed that under heat stress treatment, the addition of seaweed oligosaccharides in T1 treatment improved the effectiveness of phosphorus nutrients, and the plant growth was more fully obtained and utilized from the growth medium. The phosphorus starvation signal was low, suggesting that the effectiveness of the T1 combination nutrients was higher, which improved the plant's utilization of nutrients, obtained the best biomass, and enhanced the heat tolerance of the plant.

[0161] In general, different improved nutrient solutions will have a significant impact on the stress resistance of potato tissue culture seedlings. The improved nutrient solution with added seaweed oligosaccharides has better effects than the improved nutrient solution without additions. Among them, the T1 group has the most significant growth promotion effect and the most obvious improvement in stress resistance. After high temperature / low temperature treatment, the expression levels of stress-related enzymes and genes have significantly increased.

[0162] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. A nutrient solution for promoting potato seedling growth and enhancing stress resistance, characterized in that: The components of the nutrient solution include: 4.0-6.0 mM KNO3, 2.0-3.0 mM Ca(NO3)2, 2.0-3.0 mM NH4Cl, 0.5-1.5 mM MgSO4, 0.5-1.5 mM KH2PO4, 15.0-25.0 μM H3BO3, 4.0-5.0 μM MnCl2, 1.0-1.4 μM ZnSO4, 0.1-0.2 μM CuSO4, 0.5-1.5 mM K2SO4, 80-100 μM FeSO4, 0.1-0.2 μM (NH4)6Mo7O 24 , 0.05~0.10mM Fe-Na-EDTA.

2. The nutrient solution according to claim 1, characterized in that: The components of the nutrient solution also include 0.033-0.400 g / L of seaweed oligosaccharide.

3. The nutrient solution according to claim 1, characterized in that: The concentration of ZnSO4 in the components of the nutrient solution is 1.2 μM; The concentration of KH2PO4 in the components of the nutrient solution is 1.0 mM.

4. Use of the nutrient solution according to claim 1 in culturing potatoes.

5. A method for cultivating strong potato seedlings, characterized in that The steps include: Transplant the potato tissue culture seedlings into the growth medium and add nutrient solution for cultivation.

6. The culture method according to claim 5, characterized in that: The growth medium is a nutrient solution or a peat soil matrix; When the growth medium is peat soil matrix, the amount of nutrient solution added is twice a week, 100 mL each time.

7. The culture method according to claim 5, characterized in that: The culture time is 20 to 60 days; The culture temperature is 20-22°C.

8. The culture method according to claim 5, characterized in that: The light intensity of the culture is 5 to 15 Klux; The lighting time of the culture is 12 to 16 hours per day.

9. The culture method according to claim 5, characterized in that: The culture solution is 1 / 2 Hoagland nutrient solution.

10. The culture method according to claim 5, characterized in that: The culture solution is the nutrient solution for promoting potato seedling growth and enhancing stress resistance as described in claim 1.

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