Method for cultivating leguminous plants

Abstract

When legumes are densely cultivated in nutrient solution to increase yield in an environment where root extension is not restricted, their plant height increases, potentially leading to inefficient use of space or lodging. The technical problem of this invention is to provide a method for nutrient solution cultivation of legumes that maintains a suitable plant height for intensive cultivation. It has been found that in intensive nutrient solution cultivation at a density of 20-200 plants per square meter, using a nutrient solution with a high nitrate ion concentration can solve the aforementioned technical problem.

Description

Technical Field

[0001] This invention relates to a cultivation method for legumes using nutrient solution cultivation and to novel legume seeds. Background Technology

[0002] Plant cultivation is typically carried out through soil cultivation, which uses soil as a substrate and applies fertilizer and water as nutrients and moisture. In contrast, cultivation methods that use a nutrient solution, which is a mixture of liquid fertilizer components and water, as a substrate are called hydroponics.

[0003] In recent years, there has been a trend toward the practical application of so-called "vegetable factories," which cultivate various vegetables in factory settings. Non-patent literature 1, 2, and 3 have been published regarding research on hydroponic cultivation of soybeans. In particular, non-patent literature 1 and 3 describe a laboratory cultivation method for simultaneously cultivating multiple soybean varieties at high density. This method, which involves planting each plant in a test tube and arranging them in a tray, aims to experimentally obtain a small number of individual seeds and is not intended for practical application. Further experimental results show that both plant height and seed yield are very low.

[0004] Furthermore, Patent Document 1 discloses a method for hydroponic cultivation of legumes. The technical problem of Patent Document 1 is to provide a method for cultivating legumes that yields high-quality seeds, and it discloses a method comprising a rhizosphere low-temperature process for maintaining the roots of legumes whose seeds have already enlarged in a cooled state.

[0005] Existing technical documents

[0006] Patent documents

[0007] Patent Document 1: Japanese Patent Application Publication No. 2021-78390

[0008] Non-patent literature

[0009] Non-patent literature 1: Masao Ishimoto et al., “Exploration of genetic control factors of soybean seed components through cultivation under completely controlled conditions”, Soybean Protein Research, Vol. 19 (2016), pp. 24-29

[0010] Non-patent literature 2: Zhuang Zihe and Bo et al., “Practical application of vegetable factory (2) hydroponic cultivation conditions of edamame”, report of the Central Electric Power Research Institute, Research Report: U91033, September 1991

[0011] Non-patent literature 3: Bioscience, Biotechnology, and Biochemistry, 2015, Vol. 79, No. 1, pp. 63-67 Summary of the Invention

[0012] The problem that the invention aims to solve

[0013] When legumes are densely cultivated in nutrient solution to increase yield without restricting root extension, their plant height increases (Non-Patent Literature 2), potentially leading to inefficient use of space or lodging. The technical problem of this invention is to provide a nutrient solution cultivation method for legumes that maintains a suitable plant height for intensive cultivation. Furthermore, in other embodiments, the object of this invention is to obtain novel legume seeds, specifically edamame seeds with improved free amino acid content.

[0014] Solution for solving the problem

[0015] The inventors conducted in-depth research to solve the aforementioned technical problems. The results showed that in nutrient solution cultivation with a density of 20-200 plants per square meter, using a nutrient solution with a high nitrate ion concentration for legume cultivation can solve the above-mentioned technical problems, thus completing this invention.

[0016] That is, the present invention is as follows: (1) A method for nutrient solution cultivation of legumes, wherein a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L is used at a cultivation density of 20 to 200 plants per square meter, and cultivation is carried out in an environment that does not restrict the extension of the plant roots. (2) The cultivation method described in (1) above, wherein the cultivation method is spray cultivation or hydroponics; (3) A method for producing seeds of a legume, comprising recovering seeds from a plant obtained by means of cultivation according to (1) or (2) above; (4) A type of edamame, the amount of free asparagine is 6 to 50 mg / g on a dry basis; (5) A type of edamame, the amount of free arginine is 2.2 to 12 mg / g on a dry basis; (6) A type of edamame, wherein the amount of free asparagine is 6 to 50 mg / g on a dry matter basis and the amount of free arginine is 2.2 to 12 mg / g on a dry matter basis; (7) A method for controlling the height of leguminous plants to below 60 cm, wherein the plants are cultivated at a density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L. (8) A method for making the amount of free asparagine in edamame 6 to 50 mg / g on a dry matter basis, wherein the edamame is cultivated at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L. (9) A method for making the amount of free arginine in edamame 2.2 to 12 mg / g on a dry matter basis, wherein the edamame is cultivated at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L. (10) A method for making the amount of free asparagine in edamame 6-50 mg / g on a dry matter basis and the amount of free arginine 2.2-12 mg / g on a dry matter basis, wherein the edamame is cultivated at a cultivation density of 20-200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10-30 mmol / L. (11) A method for increasing the amount of free asparagine in edamame and mature soybean, wherein the plants are cultivated at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L.

[0017] Invention Effects

[0018] According to the present invention, a nutrient solution cultivation method can be provided for legumes that maintain a suitable plant height for nutrient solution cultivation in an intensive nutrient solution cultivation environment without restricting root extension, by using a nutrient solution with a high nitrate ion concentration. Furthermore, in other embodiments, according to the present invention, edamame with increased levels of free amino acids, particularly free asparagine and free arginine, compared to edamame grown in soil can be provided. Attached Figure Description

[0019] Figure 1 This is a diagram showing the spray cultivation device used in the embodiment.

[0020] Figure 2 This is a graph showing the plant height of soybeans cultivated in Cultivation Example 1.

[0021] Figure 3 This is a graph showing the plant height of soybeans cultivated in Cultivation Example 2.

[0022] Figure 4 This is a graph showing the yield per unit area of ​​mature soybeans cultivated in Cultivation Example 2.

[0023] Figure 5 This is a graph showing the plant height of soybeans cultivated in Cultivation Examples 3 and 4. Detailed Implementation

[0024] In one aspect of the present invention, a nutrient solution cultivation method is provided for legumes in a dense nutrient solution cultivation environment with a nitrate ion concentration of 10-30 mmol / L, using a nutrient solution in a nutrient solution with a density of 20-200 plants per square meter, in an environment that does not restrict the extension of the plant roots.

[0025] ■ Planting density

[0026] In open-field cultivation, the planting density depends on the variety, but is generally suitable at around 10-20 plants per square meter. However, in vegetable factory cultivation, higher planting density is necessary for production efficiency. Therefore, the preferred planting density for legumes in this method is 20 plants per square meter or more, more preferably 25 plants per square meter, further preferably 30 plants per square meter or more, even more preferably 35 plants per square meter or more, and still more preferably 40 plants per square meter or more. Furthermore, while increased planting density leads to increased yield, it also results in taller plants, sometimes requiring supports such as stakes. Therefore, a planting density of 200 plants per square meter or less is preferred, more preferably 150 plants per square meter or less, further preferably 120 plants per square meter or less, even more preferably 100 plants per square meter or less, and still more preferably 80 plants per square meter or less. A density of 20-200 plants per square meter is preferred, more preferably 25-150 plants per square meter, even more preferably 30-120 plants per square meter, even more preferably 35-100 plants per square meter, and still more preferably 40-80 plants per square meter.

[0027] ■Cultivation Form

[0028] The cultivation of legumes in this method is carried out in an environment that does not restrict the extension of the plant roots. That is, it is necessary to avoid spaces with liquid and / or gas phases that pre-restrict root extension, or to ensure sufficiently wide spaces. For example, it is preferable to ensure that the space for root propagation is 50 mL or more, such as 70 mL or more, 100 mL or more, 200 mL or more, 1 L or more, 5 L or more, 10 L or more, or 20 L or more. Furthermore, there is no particular upper limit, and examples include 10,000 L or less, 5,000 L or less, 2,000 L or less, 1,000 L or less, 500 L or less, 100 L or less, and 50 L or less. Each of these spaces can contain one plant or multiple plants sharing the space. It is important that root extension is not restricted; even if other plants coexist, root extension is not limited within the cultivation density range of this method. Furthermore, in one embodiment, the method of cultivation in test tubes as described in Non-Patent Documents 1 and 3 is preferably excluded.

[0029] ■Nutrient solution cultivation

[0030] The nutrient solution cultivation described in this specification refers to a cultivation method that uses a nutrient solution containing dissolved nutrients necessary for plant growth, without the use of soil. In contrast, this specification refers to cultivation methods using soil as soil cultivation. Specific examples of nutrient solution cultivation include hydroponics, solid substrate cultivation, and aeroponics. Hydroponics, as described in this specification, refers to a cultivation method in which part or all of the roots of the plant being cultivated are immersed in a nutrient solution. Solid substrate cultivation, as described in this specification, refers to a cultivation method that uses various solid substrates instead of soil. Aeroponics, as described in this specification, refers to a cultivation method in which part or all of the roots of the plant being cultivated are directly sprayed with a nutrient solution for a certain period of time. The nutrient solution cultivation of this invention refers to the aforementioned hydroponics, solid substrate cultivation, and aeroponics. The cultivation method of nutrient solution cultivation is not particularly limited. In one embodiment, the nutrient solution cultivation in this scheme is either aeroponics or hydroponics. Most preferably, hydroponics is used where the roots are fully immersed in the nutrient solution without spraying.

[0031] ■ Frequency of spraying during spray cultivation

[0032] In one embodiment, the spraying in spray cultivation is performed for 10 seconds to 1 minute at a frequency of 15 minutes to 2 hours. Preferably, it is performed at a frequency of 30 minutes to 1 hour.

[0033] ■ Nitrate ion concentration in nutrient solution

[0034] The nutrient solution in this scheme is a nutrient solution with a high nitrate ion concentration, specifically, a nutrient solution with a nitrate ion concentration of 10–30 mmol / L, for example, 10–25 mmol / L or 15–20 mmol / L. Other components such as phosphorus and potassium can be appropriately adjusted by those skilled in the art according to the legume being cultivated. For example, a nutrient solution containing various components (nitrogen, phosphorus, potassium, calcium, magnesium, sulfur, iron, manganese, etc.) in proportions suitable for general plants can be used. In a more specific embodiment, the nutrient solution in this scheme is 1 / 3 to 2 / 1 Hoagland nutrient solution. In a more specific embodiment, the nutrient solution in this scheme is Hoagland nutrient solution stock solution. Hoagland nutrient solution is known to those skilled in the art, and an example of its specific preparation method is shown below.

[0035] ■Composition of Hoagland Nutrient Solution

[0036] Stock solution H1 (dilute 200 times to 1 / 1 Hogrange solution before use)

[0037] KNO3: 101g / L (=1M)

[0038] Ca(NO3)2 4H₂O: 236 g / L (= 1 M)

[0039] Stock solution H2 (dilute 500 times to 1 / 1 Hogrange solution before use)

[0040] MgSO4 7H2O: 246g / L (=1M)

[0041] Stock solution H3 (dilute 1000 times to 1 / 1 Hogrange solution before use)

[0042] NH4H2PO4: 115g / L (=1M)

[0043] Stock solution H4 (dilute 10,000 times to 1 / 1 Hogrange solution before use)

[0044] H3BO3: 1.85 g / L (=0.03 M)

[0045] MnCl2 4H₂O: 0.99 g / L (=0.005 M)

[0046] (NH4)6Mo7O24 4H₂O: 12.36 g / L (=0.01 M)

[0047] ZnSO4 7H2O: 1.15 g / L (=0.004 M)

[0048] Stock solution Fe (dilute 10,000 times to 1 / 1 Hogrange solution before use)

[0049] Fe(III)-EDTA: 42.1 g / 500 mL (=0.2 M) should be stored in a light-proof container using aluminum foil.

[0050] Stock solution Cu (dilute 10,000 times to 1 / 1 Hogrange solution before use)

[0051] CuSO4 5H₂O: 0.25g / 500mL (=0.002M) should be stored in a light-proof container using aluminum foil.

[0052] Mix the above-mentioned stock solutions according to the following formula, and adjust the total volume with water to 5L to obtain the Hogland nutrient stock solution (15mM based on nitrate ions). H1: 25mL, H2: 10mL, H3: 5mL, H4: 500μL, Fe: 500μL, Cu: 500μL.

[0053] ■Cultivation Equipment

[0054] The cultivation method described in this scheme is typically implemented indoors. In a specific implementation, the cultivation method is carried out in equipment known as a vegetable factory. As a specific example, this equipment includes temperature control devices, light-emitting devices, nutrient solution circulation devices, and devices for adjusting carbon dioxide concentration as needed. The indoor environment described here also includes equipment that allows sunlight to pass through, such as plastic greenhouses or polytunnels. In this case, some or all of the light-emitting devices, temperature-regulating devices, and carbon dioxide concentration-regulating devices may be omitted.

[0055] ■Methods of spray cultivation

[0056] In one embodiment, in the cultivation method of this scheme, it is preferable that 50% or more, for example 80% or more, or for example 100% of the roots of the legume are exposed to the gas phase. It should be noted that exposure to the gas phase means a state where there is no direct contact with the nutrient solution, or temporary but not continuous contact via spraying or other means. 50% of the roots refers to the portion of the legume body that is equivalent to 50% by mass when the entire portion to be classified as a root is set at 100% by mass. Methods for exposing 50% or more of the roots to the gas phase include ensuring sufficient space from the lower end of the stem to the nutrient solution, and using a net or other shielding material between the roots and the nutrient solution. For standard soybeans, an example is 30 cm or more of the space to be ensured.

[0057] ■Cultivation temperature

[0058] In one embodiment, the cultivation method of this scheme is implemented by adjusting the temperature to 15–35°C, for example, 20–25°C. For example, in order to reproduce the temperature difference between day and night, the temperature can be adjusted to 25–30°C when exposed to light and to 15–20°C when not exposed to light.

[0059] ■Light irradiation device

[0060] The light irradiation device used in the cultivation method of this scheme is not particularly limited; examples include fluorescent lamps and LED lighting. LED lighting is preferred. In one embodiment, in the cultivation method of this scheme, the legume is irradiated with light at an illuminance of 5,000 to 20,000 lux, for example, 8,000 to 10,000 lux.

[0061] ■ Replacement of nutrient solution

[0062] In this cultivation method, the nutrient solution can also be used cyclically. In this case, the nutrient solution can be changed, for example, every 3 days, every 5 days, every week, every 2 weeks, or every 3 weeks. The replacement period of the nutrient solution can be determined based on the residual salt concentration. There are various methods to determine the salt concentration; for example, determining it through electrical conductivity is a straightforward method.

[0063] ■Plant height

[0064] In one embodiment, the legume plant height in the cultivation method of this scheme is less than 60 cm, for example less than 40 cm, for example 15-60 cm, 20-50 cm, 25-40 cm, 25-50 cm, or 25-60 cm. If the plant height is within this range, it has the advantages of being less prone to lodging and, in the case of vining legumes, preventing them from entangled or twining around each other. Furthermore, with this plant height, in the case of cultivation in a limited space such as a vegetable factory, the cultivation beds can be stacked vertically to increase yield, thus helping to reduce the cost of the harvest. It should be noted that, according to this scheme, even plants with relatively low plant height can achieve sufficient yields for commercial cultivation. According to this invention, which uses a nutrient solution with a nitrate ion concentration of 10-30 mmol / L and cultivates in an environment that does not restrict the extension of the plant roots, even with a cultivation density of 20-200 plants per square meter, both low plant height and high yield can be achieved.

[0065] ■Production

[0066] In one embodiment, the yield of mature seeds of the legume plant per unit area in the cultivation method of this scheme can be, for example, more than 2,000 g, more than 3,000 g, more than 4,000 g per square meter, or for example, 2,000-6,000 g, 2,500-5,000 g, or 3,000-4,500 g per square meter.

[0067] ■ Legumes

[0068] The legumes used in this cultivation method are not particularly limited and include plants broadly classified as legumes. In a more specific embodiment, the legumes are those that form root nodules when cultivated in soil. Examples of more specific legumes include adzuki beans, cowpeas, mung beans, black beans, string beans, green beans, red kidney beans, lima beans, broad beans, peas, chickpeas, lentils, soybeans, peanuts, and pigeon peas. In a more specific embodiment, the legume used in this cultivation method is soybean.

[0069] The seeds of leguminous plants mentioned in this instruction manual include both mature and immature seeds of the aforementioned plants. Furthermore, edamame refers to immature soybean seeds. More specifically, edamame refers to soybeans harvested 30–40 days after flowering, while mature soybeans refer to soybeans harvested more than 50 days after flowering.

[0070] ■ Determination of free amino acid content

[0071] The amount and ratio of free amino acids can be determined by liquid chromatography, using the following liquid chromatography-mass spectrometry method for quantification: Add 10 times (v / w) of hexane to a leguminous seed sample, such as a freeze-dried mature soybean or edamame powder sample, and mix. Remove the supernatant. Repeat the defatting and drying process three times. Then, suspend the sample in water at a concentration of 0.25% by mass, or at a concentration falling within the quantitative range of the amino acid standard solution described below, and centrifuge (14000 rpm, 5 minutes) to obtain the supernatant.

[0072] The supernatant was passed through a 0.2 μm cellulose acetate filter and separated by liquid chromatography. A Discovery HS F5 HPLC column was used, with the column temperature set to 40 °C. Solvents used were solvent A (0.1 vol% formic acid-water) and solvent B (0.1 vol% formic acid-acetonitrile). For the gradient conditions, the flow rate was set as follows: A100% to A75% / B25% for 5 min, A75% / B25% to A65% / B35% for 6 min, A65% / B35% to A5% / B95% for 4 min, A5% / B95% to A95% / B5% for 4.5 min, and A95% / B5% to A100% for 0.5 min, at a flow rate of 250 μL / min.

[0073] Samples were introduced into a mass spectrometer and analyzed in positive ion electrospray ionization mode. The analytical conditions for each amino acid are shown in Table 1. Amino acid mixed standard solution H type (Fujifilm and Kojun Pharmaceutical Co., Ltd.) can be used as a standard for quantification.

[0074] [Table 1]

[0075] ■ Amount of free amino acids in edamame

[0076] In one embodiment, compared to edamame obtained through soil cultivation or ordinary hydroponics, the edamame obtained through the cultivation method of this scheme has an increased amount of free amino acids. In a more specific embodiment, the amount of free amino acids in edamame obtained through the cultivation method of this scheme, on a dry matter basis, can be 20 mg / g to 100 mg / g, for example 25 mg / g to 95 mg / g, or 30 mg / g to 90 mg / g. In a more specific embodiment, the amount of free asparagine in edamame obtained through the cultivation method of this scheme, on a dry matter basis, can be 6.0 mg / g to 50 mg / g, for example 8.0 mg / g to 45 mg / g, or 10 mg / g to 40 mg / g. In a more specific embodiment, the amount of free serine in edamame obtained by the cultivation method of this scheme, on a dry matter basis, can be 0.40 mg / g to 5.0 mg / g, for example 0.50 mg / g to 4.0 mg / g, or 0.60 mg / g to 3.0 mg / g. In a more specific embodiment, the amount of free alanine in edamame obtained by the cultivation method of this scheme, on a dry matter basis, can be 1.0 to 25 mg / g, for example 2.0 to 20 mg / g, or 3.0 to 15 mg / g. In a more specific embodiment, the amount of free glutamine in edamame obtained by the cultivation method of this scheme, on a dry matter basis, can be 0.25 to 10 mg / g, for example 0.40 to 8.0 mg / g, 0.60 to 7.0 mg / g, or 0.80 to 6.0 mg / g. In a more specific embodiment, the amount of free histidine in edamame obtained by the cultivation method of this scheme, calculated on a dry matter basis, can be 1.2–12 mg / g, for example 1.5–10 mg / g, or 2.0–8.0 mg / g. In a more specific embodiment, the amount of free arginine in edamame obtained by the cultivation method of this scheme, calculated on a dry matter basis, can be 2.2–12 mg / g, for example 2.7–10 mg / g, or 3.2–8.0 mg / g. In a more specific embodiment, the amount of free threonine in edamame obtained by the cultivation method of this scheme, calculated on a dry matter basis, can be 0.45–4.0 mg / g, for example 0.55–3.0 mg / g, or 0.70–2.0 mg / g.

[0077] Therefore, in one embodiment, the present invention provides edamame with the aforementioned amount of free amino acids.

[0078] In one embodiment, mature soybeans obtained through the cultivation method of this scheme have an increased amount of free amino acids compared to mature soybeans obtained through soil cultivation or ordinary hydroponics. In a more specific embodiment, the amount of free amino acids in mature soybeans obtained through the cultivation method of this scheme, calculated on a dry matter basis, can be 3.0–20 mg / g, for example, 3.5–15 mg / g or 5.0–15 mg / g. In a more specific embodiment, the amount of free asparagine in mature soybeans obtained through the cultivation method of this scheme, calculated on a dry matter basis, can be 0.30–10 mg / g, 0.50–7.0 mg / g or 0.80–5.0 mg / g.

[0079] Therefore, in one embodiment, the present invention provides mature soybeans with the aforementioned amount of free amino acids.

[0080] ■ The effect of increasing free amino acids

[0081] The free asparagine content of the aforementioned edamame is increased compared to previous edamame, which is expected to improve endurance and fatigue recovery. Furthermore, the free arginine content is also increased, similarly expected to improve fatigue recovery. Additionally, the histidine content is increased, which is expected to improve concentration and memory. Furthermore, the threonine content is increased, which is expected to improve gastritis. Furthermore, the glutamine content is increased, which is expected to accelerate muscle fatigue recovery. Furthermore, the alanine content is increased, which is expected to promote fat burning. Furthermore, the serine content is increased, which is expected to moisturize the skin. Moreover, the free asparagine content of the aforementioned mature soybeans is increased compared to existing mature soybeans, which is expected to improve endurance and fatigue recovery. In specific embodiments, the aforementioned edamame or mature soybeans can be used as raw materials for supplements, etc.

[0082] In one embodiment, the present invention provides a method for manufacturing seeds of a legume, comprising recovering seeds from a legume obtained through the above-described cultivation method. Furthermore, in other embodiments, the present invention provides offspring obtained from seeds obtained through the above-described manufacturing method.

[0083] In the above-mentioned schemes for immature seeds, mature seeds, manufacturing methods, and progeny, all the matters described in the above-mentioned cultivation method schemes apply to these schemes.

[0084] Example

[0085] The following examples illustrate the implementation of the present invention in more detail. It should be noted that, unless otherwise specified, "%" and other terms in the examples are based on a mass standard.

[0086] Cultivation Example 1: A Study on Nutrient Solution Spray Cultivation of Soybeans

[0087] For soybean varieties, Kamikazeka, Mifuka, Sayamasume, and Yukine (all from Snow Brand Seedling Co., Ltd.) were used. For the spray cultivation system, Sodatec Aero (Yakumo Trading Co., Ltd.) was used. An overview of the spray cultivation system is shown below. Figure 1 Mature soybeans of various varieties were germinated on absorbent seedling paper and transplanted into a spray culture device when the hypocotyl reached approximately 5 cm in length. The chamber dimensions were 60 cm × 40 cm × 18 cm, the pump discharge pressure was 80 psi, and the pump flow rate was 0.85 L / min. Hoagland nutrient solution (15 mM nitrate ion concentration) was used as the nutrient solution. The planting density was 25 plants per square meter. The root extension space was approximately 43 L. In the spray culture device, the nutrient solution was sprayed directly onto the roots for 20 seconds every 30 minutes. The nutrient solution was changed every week. Irradiation was provided 13 times daily with LED lighting at approximately 10,000 lux, with the temperature adjusted to 25°C during illumination and 20°C during non-illumination periods. It should be noted that a mesh was placed 1 cm above the nutrient solution surface at the bottom of the chamber to ensure that more than 50% of the roots were exposed to the gas phase.

[0088] When using a compact conductivity meter (LAQUAtwin EC-33B) When the conductivity of the nutrient solution was measured (using HORIBA), it was 2.3 mS / cm immediately after the nutrient solution was changed, and 2.2 mS / cm one week later; no significant reduction in salts was observed. Furthermore, no large root nodules were observed in this cultivation method. Each variety flowered 31–32 days after sowing. Edamame can be harvested approximately 30–40 days after flowering, and mature soybeans approximately 60–70 days later. The plant height of all varieties was 30–40 cm, within the target range (…). Figure 2 In addition, the taproot and most of the lateral roots remain in the gas phase on the upper surface of the net. It should be noted that the yield per square meter is 2346–2644 g.

[0089] Cultivation Example 2: A Study on Cultivation Density

[0090] Under the same conditions as in Cultivation Example 1, the planting density was adjusted to 25, 50, 75, and 100 plants per square meter, and cultivation was carried out accordingly. Although a tendency was observed that higher planting density resulted in taller plants, in this cultivation example using a nutrient solution with a high nitrate ion concentration, the plant height was consistently below 60 cm, falling within the target range. Furthermore, the plant heights at 25, 50, and 75 plants per square meter were all below 40 cm. Figure 3 ).

[0091] The yield of mature soybeans per unit area is shown in the figure. Figure 4There is no difference in yield among varieties. As planting density increases, yield increases, but the slope gradually flattens out.

[0092] Cultivation Example 3: Hydroponic Cultivation (Hogland Solution / 15mM Nitrate Ions)

[0093] Soybean hydroponics was conducted under conditions where the roots were not exposed to the gas phase. A HomeHyponica MUSUCO hydroponic system was used as the hydroponic system. Eight plants (50 plants per square meter) were cultivated in a 40cm × 39.6cm × 26.3cm device under the same nutrient solution and irradiation conditions as in Example 1. The root extension space was approximately 42L. 15L of nutrient solution was stored in a liquid fertilizer tank and circulated, being replaced every week.

[0094] In this cultivation method, no soybean root nodules were observed. Edamame can be harvested approximately 30-40 days after flowering, and mature soybeans can be harvested approximately 60-70 days later. The plant height for any variety is in the range of 30-40 cm. Figure 5 In addition, the yield per square meter is 2766–4093 g.

[0095] Cultivation Example 4: Hydroponic cultivation (typically nutrient solution / nitrate ion 7.5mM)

[0096] In a nutrient solution with a typical concentration of 7.5 mM nitrate ions, intensive cultivation was carried out at a density of 25 plants per square meter. The hydroponic cultivation apparatus used was the same as that used in Cultivation Example 3. The root extension space was approximately 42 L. Cultivation was carried out according to Cultivation Example 1, using the following nutrient solution: Total nitrogen 115 (same as below ppm), ammonia nitrogen 10, nitrate nitrogen 105, phosphate (P2O5) 46.5, potassium (K2O) 188.5, calcium (CaO) 109.5, magnesium (MgO) 40, manganese (MnO) 0.5, boron (B2O3) 0.5, iron (Fe) 1.45, copper (Cu) 0.01, zinc (Zn) 0.02, molybdenum (Mo) 0.01.

[0097] The plant height of any variety ranges from 57 to 74 cm. Figure 5 In this cultivation example, where a nutrient solution with a typical nitrate ion concentration was used, plant heights exceeding 60 cm were also observed. Plants exceeding 60 cm in height were susceptible to lodging. Furthermore, the yield per square meter ranged from 811 to 1966 g.

[0098] Cultivation Example 5: Ultra-high density test tube culture

[0099] An experiment was conducted with reference to Non-Patent Document 3. A 12-well test tube rack (“4-WAY Flipper”) was used. Two 17cm x 5cm tubes (INA-OPTIKA type) were connected together as a group, with the top surface of each tube being the same. Six 12ml culture tubes (INA-OPTIKA type), each with two openings 1cm and 2cm from the bottom, were inserted alternately into the previously prepared 24-well test tube rack and placed on a tray for cultivation. The soybean variety used was Yukino. Other conditions followed those in Cultivation Example 1. The root space was approximately 12mL. The cultivation density in this experiment was approximately 353 plants per square meter. Furthermore, the average plant height was 20.9cm, which was very low.

[0100] In other cultivation examples in this manual, the fruit yield was over 30g / plant, while in contrast, this cultivation example only yielded 2.32g / plant, indicating significantly insufficient growth.

[0101] Cultivation Example 6: High-density test tube culture

[0102] Referring to Non-Patent Literature 1, cultivation was conducted in the same manner as in Cultivation Example 5. A 12 mL culture tube (made by INA-OPTIKA) was inserted into the previously prepared 24-well test tube rack and placed on a tray for cultivation. The soybean variety used was Kamikaze. Other conditions were the same as in Cultivation Example 1. The root extension space was approximately 12 mL. The cultivation density in this experiment was approximately 59 plants per square meter. Furthermore, the average plant height was 21.5 cm, which was very low.

[0103] The yield of this cultivation example remained at 2.39 g / plant, showing significantly insufficient growth, similar to cultivation example 5.

[0104] Based on cultivation examples 5 and 6, it is believed that in cultivation methods where the root extension range is restricted, such as in test tubes, the plant cannot adequately obtain the nutrients required for growth, regardless of the cultivation density, resulting in insufficient growth. These cultivation methods, as described above, are merely experimental techniques for obtaining small quantities of seeds. Even with these existing technological insights, it is impossible to conceive of solutions to the influence of nitrate ion concentration on plant height, or even the technical problems of plant height extension in high-density cultivation.

[0105] Cultivation Example 7: Study on the Content of Free Amino Acids

[0106] The free amino acid content in edamame harvested 31–36 days after flowering and mature soybeans harvested 60–70 days after flowering of various varieties was determined. The results of cultivation example 1, grown using nutrient solution spraying, are shown in Table 2, and the results of cultivation example 3, grown using hydroponics, are shown in Table 3. For comparison, the results of cultivation example 4, grown using conventional nutrient solution hydroponics, are shown in Table 4, and the free amino acid content in edamame and mature soybeans obtained through outdoor soil cultivation is shown in Table 5.

[0107] ■Edamame and mature soybeans harvested through nutrient solution spraying with a nutrient solution of high nitrate ion concentration. Free amino acid content in cooked soybeans (mature beans)

[0108] [Table 2]

[0109] ■Edamame and mature soybeans harvested using hydroponics with a high concentration of nitrate ions. Free amino acid content in (mature beans)

[0110] [Table 3]

[0111] ■Free edamame and mature soybeans (mature beans) harvested through hydroponics using conventional nutrient solutions amino acid content

[0112] [Table 4]

[0113] ■ Free amino acid content in edamame and mature soybeans (mature beans) harvested through soil cultivation

[0114] [Table 5]

[0115] The results in Tables 2 (nutrient solution spraying) and 3 (hydroponics) compared to Table 4 (normal nutrient solution hydroponics) showed an increase in the amount of free amino acids in edamame and mature soybeans. In edamame, the amounts of free asparagine, serine, alanine, glutamine, threonine, histidine, and arginine were particularly increased. In mature soybeans, the amount of free asparagine was particularly increased. Furthermore, Tables 2 and 3 compared to Table 5 (soil cultivation) showed an increase in the amount of free amino acids in edamame and mature soybeans. In edamame, the amounts of free asparagine, serine, alanine, glutamine, glutamic acid, histidine, and arginine were particularly increased. In mature soybeans, the amount of free asparagine was particularly increased.

[0116] When edamame grown in nutrient solution spraying (Cultivation Example 1), hydroponics (Cultivation Example 3), conventional nutrient solution hydroponics (Cultivation Example 4), and soil cultivation were tasted, regardless of the variety, the spray-grown and hydroponically grown legumes had a richer flavor compared to the conventional nutrient solution hydroponics and soil cultivation legumes.

[0117] Industrial availability

[0118] This invention provides a method for maintaining a suitable plant height for nutrient solution cultivation in legumes under high planting density. Furthermore, this invention also provides edamame with increased levels of free asparagine and free arginine. This invention can be applied in a wide range of fields, including agriculture and food production.

Claims

1. A method for hydroponic cultivation of legumes, wherein, Cultivate at a planting density of 20 to 200 plants per square meter, using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L, in an environment that does not restrict the extension of the plant roots.

2. The cultivation method according to claim 1, wherein, The cultivation method is either spray cultivation or hydroponics.

3. A method for producing seeds of a legume, comprising recovering seeds from a plant obtained by the cultivation method according to claim 1 or 2.

4. A type of edamame, wherein the amount of free asparagine is 6-50 mg / g on a dry weight basis.

5. A type of edamame, wherein the amount of free arginine, calculated on a dry matter basis, is 2.2–12 mg / g.

6. A type of edamame, wherein the amount of free asparagine is 6-50 mg / g on a dry matter basis, and the amount of free arginine is 2.2-12 mg / g on a dry matter basis.

7. A method for controlling the plant height of leguminous plants to below 60 cm, wherein, Cultivate at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L.

8. A method for reducing the amount of free asparagine in edamame to 6-50 mg / g on a dry matter basis, wherein, Cultivate at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L.

9. A method for reducing the amount of free arginine in edamame to 2.2–12 mg / g on a dry matter basis, wherein, Cultivate at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L.

10. A method for reducing the amount of free asparagine in edamame to 6-50 mg / g on a dry matter basis and the amount of free arginine to 2.2-12 mg / g on a dry matter basis, wherein... Cultivate at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L.

11. A method for increasing the amount of free asparagine in edamame and mature soybeans, wherein, Cultivate at a planting density of 20 to 200 plants per square meter using a nutrient solution with a nitrate ion concentration of 10 to 30 mmol / L.

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