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WO2021112139A1 - Promoteur de formation de nodules de pois chiche - Google Patents

Promoteur de formation de nodules de pois chiche Download PDF

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Publication number
WO2021112139A1
WO2021112139A1 PCT/JP2020/044924 JP2020044924W WO2021112139A1 WO 2021112139 A1 WO2021112139 A1 WO 2021112139A1 JP 2020044924 W JP2020044924 W JP 2020044924W WO 2021112139 A1 WO2021112139 A1 WO 2021112139A1
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Prior art keywords
chickpea
rhizobia
soil
methylated
isoflavones
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English (en)
Japanese (ja)
Inventor
雄平 津野
厚輝 大西
輝久 藤松
遠藤　圭二
春香 前田
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Kao Corp
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Kao Corp
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Priority to AU2020395007A priority Critical patent/AU2020395007B2/en
Publication of WO2021112139A1 publication Critical patent/WO2021112139A1/fr
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    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G7/00—Botany in general
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G7/00—Botany in general
    • A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N43/00—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds
    • A01N43/02—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms
    • A01N43/04—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom
    • A01N43/14—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom six-membered rings
    • A01N43/16—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom six-membered rings with oxygen as the ring hetero atom
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
    • C05F11/00—Other organic fertilisers
    • C05F11/10—Fertilisers containing plant vitamins or hormones

Definitions

  • the present invention relates to a technique for promoting nodule formation in chickpea and increasing revenue of chickpea.
  • Chickpea (Cicer arietinum L.) is an annual herb of the leguminous family, and grains (beans), young foliage and pods are edible. Dried chickpea seeds are rich in protein 25%, fat 5%, carbohydrate 59%, dietary fiber 1.2%, and minerals such as phosphorus, calcium, and iron. This has the effect of refraining from animal protein in diets and the like and supplementing essential amino acids that tend to be deficient in grains (Non-Patent Document 1).
  • Chickpea occupies the second place in the harvested area and the third place in the production volume among the legumes (Pulses) in the whole world, and is the legume with the highest production volume and demand in India where the population is increasing rapidly. However, it is said that the yield of chickpeas in India is different between the test field level and the actual producer level (Non-Patent Document 2).
  • Non-Patent Document 3 It is known that the yield of chickpea increases due to nodule formation by rhizobia (Mesorhizobium ciceri, etc.) (Non-Patent Document 3), and it is considered important to enhance the nodule to ensure productivity.
  • Conventionally, techniques such as adding a specific compound to proliferate and activate rhizobia have been reported, but chickworms have been empirically used to activate rhizobia that coexist with soybeans and common beans. Flavonoids such as daidzein and naringenin are used, and their effects are not sufficient.
  • methylated isoflavones such as biochanin A and formononetin, which are abundant in red clover, have long been known to have a female hormone-like action, and recently, they have a dementia-relieving effect (Patent Document 1) and sarcopenia. It has been reported that there is a preventive or therapeutic effect (Patent Document 2). However, it is not known at all what kind of action methylated isoflavones have on root nodules.
  • Patent Document 1 Japanese Patent Application Laid-Open No. 2019-526612
  • Patent Document 2 Japanese Patent Application Laid-Open No. 2013-507361
  • Non-Patent Document 1 Coastal Bioenvironment Center Saga University Volume 8 (2006) 35-40
  • Non-Patent Document 2 Outlook on Agriculture, Volume 46, Issue 3 (2017) 1-6
  • Non-Patent Document 3 European Journal of Agronomy, Volume 19, Issue 1 (2003) 15-21
  • the present invention relates to the following 1) to 8).
  • a method for promoting nodule formation of chickpea which comprises a step of applying methylated isoflavones represented by the following formula (1) to soil or plants.
  • a method for increasing the yield of chickpea which comprises a step of applying methylated isoflavones represented by the following formula (1) to soil or plants.
  • a method for cultivating chickpeas which comprises a step of applying methylated isoflavones represented by the following formula (1) to soil or plants.
  • R 1 and R 2 may be the same or different and represent a hydrogen atom or a hydroxy group.
  • Nodule formation promoting effect of methylated isoflavones nodule weight. Nodule formation promoting effect of methylated isoflavones (number of nodules). Initial growth promoting effect of methylated isoflavones (aboveground weight). Initial growth promoting effect of methylated isoflavones (underground weight). Yield increasing effect of methylated isoflavones (grain weight). Yield increasing effect of methylated isoflavones (real number). Synergistic effect (number of nodules) by the combined use of rhizobia and prtensein. Synergistic effect of combined use of rhizobia and prtensein (nodule weight).
  • the present invention relates to providing a technique for promoting nodule formation by rhizobia coexisting with chickpea and increasing the yield of chickpea.
  • root nodule formation by rhizobia symbiotic with chickpea is promoted, and the yield of chickpea can be increased.
  • the term “chickpea” refers to Cicers arietinum L. et al. Of the genus Cicers in the family Leguminosae. Means. As the species, fog species and desi species are known, but any of them may be used.
  • the "root nodule” means a nodule formed in the root of a plant by coexistence with a bacterium (rhizobia). Rhizobia perform so-called symbiotic nitrogen fixation that reduces atmospheric nitrogen in the nodules to convert it into ammonia nitrogen and supplies it to the host.
  • Rhizobia are said to have specificity for symbiotic plants, and Mesorhizobium medianeum and the like are known as rhizobia that use chickworm as a host. Has been done.
  • the "rhizobium” is not limited as long as it can coexist with chickpea, but is preferably Mesorhizobium ciselli.
  • the methylated isoflavones are isoflavones in which the hydroxy group at the 4'position is methylated, as represented by the following formula (1).
  • R 1 and R 2 may be the same or different and represent a hydrogen atom or a hydroxy group.
  • a compound in which both R 1 and R 2 are hydroxy groups (5,7,3'-trihydroxy-4'-methoxyisoflavone) is known as "pratensein", and R 1 is a hydroxy group.
  • the compound in which R 2 is a hydrogen atom (5,7-dihydroxy-4'-methoxyisoflavon) is known as “Biochanin A”, and the compound in which both R 1 and R 2 are hydrogen atoms (5,7-dihydroxy-4'-methoxyisoflavon).
  • 7-Hydroxy-4'-methoxyisoflavone) is known as "Formonetin".
  • prtensein is preferable from the viewpoint of promoting nodule formation.
  • Such methylated isoflavones of the present invention may be contained in plants such as red clover (red clover, Trifolium pratense), white clover (white clover, Trifolium repens), nasbitohagi (Podocarpium podocarpum), and hariganegoke (Bryum capillare). It has been reported, and generally, it can be extracted, isolated, and purified from the plant by a known method, but commercially available products (Platinesin: Fujifilm Wako Junyakusha, Chromadex; Biochanin A: Chromadex; Formononetin: Sigma-Aldrich) can also be used. Also, by a previously reported report (for example, Wong, E. 1963. “Pratensein. 5, 7, 3 ′-Trihydroxy-4 ′-Methoxyisoflavone.” Journal of Organic Chemistry 28 (9): 2336-39.) It may be a manufactured chemically synthesized product.
  • the addition of the methylated isoflavones of the present invention increases the nodule weight and the number of nodules, resulting in an individual plant. It increases nodule weight, number of nodules and protein yield per nodule, and its action is superior to that of naringenin (4', 5,7-trihydroxyflavanone).
  • the methylated isoflavones of the present invention can serve as a nodule formation promoter for chickpeas or a nodule forming agent for chickpeas, and can be used for promoting nodule formation of chickpeas or increasing the yield of chickpeas, and root nodules of chickpeas. It can be used to produce nodule promoters or chickpea yield enhancers.
  • promoting nodule formation of chickpea means that the ability of rhizobia to form nodules in symbiosis with chickpea is increased, and specifically, the nodule weight, the number of nodules, or both are methyl. It means that it increases more than the untreated group of isoflavones.
  • increased yield of chickpea means that the number of grains (beans) harvested from chickpea increases, for example, the dried grain weight and / or the number of grains per plant are methylated isoflavones. It means that it increases more than the untreated group.
  • “Increased chickpea yield” also includes an increase in the protein yield from chickpeas.
  • “Increased protein yield” means that the mass of protein contained in grains (beans) harvested from chickpeas increases, for example, the mass of protein contained in grains (beans) per individual plant is methyl. It means that it increases more than the untreated group of isoflavones.
  • the above-mentioned chickpea nodule formation promoter or yield enhancer is a composition for promoting nodule formation or yield increase of chickpea (composition for promoting nodule formation or yield increase of chickpea) (for example, various agricultural or horticultural materials). ), Or it can be a material (single unit) or a preparation for adding or blending chickpeas directly or diluted with water to a cultivation base material for cultivating chickpeas such as soil, medium, and nutrient solution cultivation solution.
  • the form of the composition may be a liquid or gel-like composition, or may be a solid state (block-like, powder-like, granular, etc.) composition.
  • a liquid composition it can be a concentrated type that is used as it is or after being diluted. Further, in the case of a composition in a solid state, it can be used by dissolving it in water.
  • the above composition may contain any component in addition to the methylated isoflavones of the present invention.
  • Such components include solvents (eg, water, buffers, media, hydroponic cultivation solutions, etc.), carriers (diatomaceous soil, vermiculite, pearlite, peat moss, activated charcoal, humanus, talc, zeolite, clay, carbon black, pulp.
  • Plant growth promoting microorganisms such as and mycorrhizal fungi, essential nutrients of plants, flavonoids (dizein, genistein, etc.), organic acids, amino acids, peptides, nucleosides, nucleotides, nucleic acid bases, sugars, monovalent alcohols, nonionic surfactants , Food additives, microbial extracts, plant hormones, nod factors ie lipo-chito-oligosaccharides, synthetic lipo-chito-oligosaccharides, chito-oligosaccharides, chitinic compounds, linoleic acid or its derivatives, linolenic acid or its derivatives, calikin, acyl -Homoserine lactone derivatives, betaine compounds, phenolic compounds (ferulic acid, chlorogenic acid, etc.), antioxidants and the like can be mentioned.
  • Examples of the above composition include, for example, a cultivation base material containing the methylated isoflavones of the present invention as an active ingredient (for example, soil for agriculture or horticulture, soil, medium, solution for nutrient solution cultivation, water, etc.).
  • a cultivation base material containing the methylated isoflavones of the present invention as an active ingredient for example, soil for agriculture or horticulture, soil, medium, solution for nutrient solution cultivation, water, etc.
  • Examples include fertilizers, water for watering, microbial materials such as rhizobial materials, soil conditioners, pesticides, seeding materials, plant supplements (for example, activators, nutrients, etc.), but are limited to these. Not done.
  • the fertilizer, microbial material, soil conditioner, sowing material, and plant supplement are preferable because they contribute to the improvement of the soil for cultivating plants such as legumes.
  • the fertilizer, microbial material, soil conditioner, sowing material, plant supplement may be solid or liquid, and in the case of solid, it may be block-like, powder-like, granular or the like, but powder. Alternatively, it is preferably granules.
  • the fertilizer, microbial material, soil conditioner, sowing material, and plant supplement contain the methylated isoflavones of the present invention as an active ingredient, and the fertilizer, microbial material, and soil conditioner usually used for chickpea cultivation. It may contain ingredients of agents, seeding materials, plant supplements.
  • the cultivated base material, fertilizer, microbial material such as rhizobial material, soil improver, pesticide, sowing material, plant supplement can be used as a normal cultivated base material (for example, soil for agriculture or gardening, soil, medium, etc. Hydroponic cultivation solution, water, etc.), fertilizers, microbial materials such as rhizobial materials, soil conditioners, pesticides, seeding materials, plant supplements (eg, activators, nutrients, etc.), etc. It may be prepared by adding methylated isoflavones.
  • the content of the methylated isoflavones of the present invention in the composition for promoting root grain formation or increasing the yield of chicks can be appropriately set so as to be suitable for the application rate, and for example, it is preferably in the total mass of the composition.
  • 0.000001% by mass (0.01 ⁇ g / g) or more, more preferably 0.00001% by mass (0.1 ⁇ g / g) or more, still more preferably 0.0001% by mass (1 ⁇ g / g) or more, and preferably Is 1% by mass (10,000 ⁇ g / g) or less, more preferably 0.1% by mass (1,000 ⁇ g / g or less, still more preferably 0.01% by mass (100 ⁇ g / g) or less, still more preferably 0. It is 001% by mass (10 ⁇ g / g) or less, preferably 0.000001 to 1% by mass, more preferably 0.00001 to 0.1% by mass, still more preferably 0.0001 to 0.01% by mass. Even more preferably, it is 0.0001 to 0.001% by mass.
  • the composition for promoting nodule formation or increasing yield of chicks includes the methylated isoflavones of the present invention and rhizobia using chicks as hosts (for example, Mesorhizobium ciselli, Mesorhizobium meditera) from the viewpoint of promoting root nodule formation of chicks.
  • a composition that is a microbial material containing Mesorhizobium (genus Mesorhizobium root nodule bacteria) such as neum is preferable, and a material containing Mesorhizobium cyseri is particularly preferable.
  • Cell count of the rhizobia can be set appropriately to suit the application rate, for example, composition per 1g, the number of rhizobia preferably 10 4 cfu or more, more preferably 10 5 cfu or more, more preferably Is 10 6 cfu or more, and preferably 10 11 cfu or less, more preferably 10 10 cfu or less, still more preferably 10 9 cfu or less, still more preferably 5 ⁇ 10 8 cfu or less. Further, it is preferably 10 4 to 10 11 cfu, more preferably 10 5 to 10 10 cfu, still more preferably 10 6 to 10 9 cfu, and even more preferably 2 ⁇ 10 6 to 5 ⁇ 10 8 cfu.
  • the content of rhizobia and methylation isoflavones for example, composition per 1g, preferably the number of rhizobia 10 4 ⁇ 10 11 cfu and methylation isoflavones 0.01 ⁇ 10,000 [, more preferably Rhizobia is 10 5 to 10 10 cfu and methylated isoflavones are 0.1 to 1,000 ⁇ g, more preferably rhizobia is 10 6 to 10 9 cfu and methylated isoflavones are 1 to 100 ⁇ g, and even more preferably root grains. bacteria 2 ⁇ 10 6 ⁇ 5 ⁇ 10 8 cfu and methylation isoflavones is 1 ⁇ 10 [mu] g.
  • the content ratio of rhizobia and methylated isoflavones is preferably from 10 5 ⁇ 5 ⁇ 10 9, more preferably 10 6 ⁇ 5 ⁇ 10 9, more preferably Is 2 ⁇ 10 6 to 5 ⁇ 10 9 , and even more preferably 2 ⁇ 10 6 to 5 ⁇ 10 8 .
  • the method for supplying the nodule formation promoting agent or the yield increasing agent for chickpeas of the present invention is not particularly limited as long as it is applied so as to promote the nodule formation in chickpeas and exert the effect of increasing the yield of chickpeas. That is, as long as the root grain formation promoter or yield enhancer of the chick of the present invention comes into contact with or is delivered to the soil of the plant body or the rhizosphere of the plant, there is no particular limitation, and surface spraying, irrigation, or digging into the soil. , Foliar spraying on plants, application mixed with fertilizer, addition to hydroponic solution, application or smearing on seeds before sowing (for example, seed powder coating) and the like.
  • the soil a soil capable of cultivating chickpea may be used, and a soil to which a soil fungus solution prepared from a soil capable of cultivating chickpea is added can also be used.
  • the soil bacterial solution can be prepared as a filtrate obtained by filtering the soil to which sterilized water having a mass of 0.1 to 10 times the mass of the soil has been added.
  • rhizobia for example, a solution containing rhizobia
  • methylated isoflavones are added to the soil in which the seeds of the plant are sown, respectively, or the above-mentioned rhizobia using chickpea as a host and methylation of the present invention are added.
  • examples include preparing a composition (microbial material) containing isoflavones and adding the material to chickpea seeds or soil in which chickpeas are cultivated.
  • the application rate of the root grain formation promoter or yield enhancer of the chick of the present invention may differ depending on the application method, application time, plant type, cultivation density, growth stage, etc., but for example, as the methylated isoflavones of the present invention.
  • the amount of syrup used is preferably 0.000001 to 1 mass ppm, more preferably 0.00001 to 0.1 mass ppm, still more preferably 0.0001 to 0.0001 to 1 mass ppm as the concentration in the cultivation base material for cultivating chicks. It may be 0.01 mass ppm, more preferably 0.0001 to 0.001 mass ppm.
  • the amount of the methylated isoflavones of the present invention used per 1 liter volume of the cultivation base material is preferably 0.001 to 1,000 ⁇ g, more preferably 0.01 to 100 ⁇ g, still more preferably 0.1 to 10 ⁇ g. Even more preferably, it may be 0.1 to 1 ⁇ g.
  • chickpeas are cultivated in soil, preferably 0.0001 to 100 g, more preferably 0.001 to 10 g, still more preferably 0.01 to 1 g, and even more preferably 0.01 to per 10 ares of land.
  • the methylated isoflavones of the present invention may be added to the soil in an amount of 0.1 g.
  • the amount of the composition used is the composition. It depends on the concentration of methylated isoflavones contained therein. For example, when the concentration of the methylated isoflavones contained in the composition is 0.005% by mass, the amount of the composition used per 10 ares of land is preferably 2 to 2,000,000 g. It is preferably 20 to 200,000 g, more preferably 200 to 20,000 g, and even more preferably 200 to 2,000 g.
  • the nodule formation promoter or yield increasing agent for chickpea of the present invention may be applied in an amount in the above range at a time, or may be applied in a plurality of times.
  • the number of rhizobia to be applied is one seed of chicks.
  • the application rate of the methylated isoflavones in the combination application is preferably 0.001 ⁇ g or more, more preferably 0.01 ⁇ g or more, still more preferably 0.1 ⁇ g or more, and preferably 0.1 ⁇ g or more per chickpea seed. It is 1,000 ⁇ g or less, more preferably 100 ⁇ g or less, still more preferably 10 ⁇ g or less, still more preferably 5 ⁇ g or less.
  • the dose per one grain seeds chickpea rhizobia and methylated isoflavones in the combination application preferably is rhizobia 10 4 ⁇ 10 11 cfu and methylation isoflavones 0.001 ⁇ 1,000 .mu.g, more preferably the rhizobia 10 5 ⁇ 10 10 cfu and methylation isoflavones 0.01 ⁇ 100 [mu] g, more preferably rhizobia is 10 6 ⁇ 10 9 cfu and methylation isoflavones 0.1 ⁇ 10 [mu] g, still more Rhizobia is preferably 2 ⁇ 10 6 to 5 ⁇ 10 8 cfu and methylated isoflavones are 0.1 to 5 ⁇ g.
  • the application rates of ratio of rhizobia and methylated isoflavones in the combination application is preferably 10 5 ⁇ 5 ⁇ 10 9, more preferably 10 6 ⁇ 5 ⁇ 10 9, more preferably 2 ⁇ 10 6 ⁇ 5 ⁇ 10 9, even more preferably from 2 ⁇ 10 6 ⁇ 5 ⁇ 10 8.
  • the timing and frequency of application of the chickpea root grain formation promoter or yield enhancer may vary depending on the type of plant, etc., but is usually surface spraying, irrigation, plow-in, or seed powder coating on a cultivation base material such as soil.
  • a cultivation base material such as soil.
  • it is preferably applied once or 1 to 3 times between the day of sowing and 30 days, preferably between the day of sowing and 14 days.
  • the chickpea of the present invention is applied to a cultivation base material (for example, soil, soil, medium, solution for hydroponic cultivation, water, etc.) before sowing, at the same time as sowing, and / or after sowing.
  • a cultivation base material for example, soil, soil, medium, solution for hydroponic cultivation, water, etc.
  • the addition of the root grain formation promoter or yield enhancer may be combined with the application or smearing (for example, seed powder coating) of the chickpea root grain formation promoter or yield enhancer to the seeds before sowing.
  • a carrier such as peat moss, a spreading agent, a surfactant or the like is used. Is preferable.
  • a nodule formation promoter for chickpeas containing methylated isoflavones represented by the following formula (1) as an active ingredient ⁇ 2> An agent for increasing the yield of chickpeas containing methylated isoflavones represented by the following formula (1) as an active ingredient.
  • a composition for promoting nodule formation or increasing yield of chickpeas containing methylated isoflavones represented by the following formula (1) and rhizobia using chickpeas as hosts ⁇ 3> A composition for promoting nodule formation or increasing yield of chickpeas containing methylated isoflavones represented by the following formula (1) and rhizobia using chickpeas as hosts.
  • ⁇ 4> A method for promoting nodule formation of chickpea, which comprises a step of applying methylated isoflavones represented by the following formula (1) to soil or plants.
  • ⁇ 5> A method for increasing the yield of chickpea, which comprises a step of applying methylated isoflavones represented by the following formula (1) to soil or plants.
  • ⁇ 6> A method for cultivating chickpeas, which comprises a step of applying methylated isoflavones represented by the following formula (1) to soil or plants.
  • ⁇ 7> The method according to any one of ⁇ 4> to ⁇ 6>, wherein the soil or plant is further applied in combination with rhizobia having chickpea as a host.
  • ⁇ 8> The method according to ⁇ 7>, wherein a microbial material containing rhizobia and methylated isoflavones is prepared in advance, and the material is applied to chickpea seeds or soil in which chickpeas are cultivated.
  • the root nodule bacterium is a root nodule bacterium of the genus Mesorhizobium (genus Mesorhizobium), preferably Mesorhizobium ciceri or Mesorhizobium mediterraneum (described in Mesorhizobium media7 or ⁇ method> 8).
  • the soil is a soil to which a soil fungus solution prepared from a soil capable of cultivating chickpea is added.
  • ⁇ 11> Use of methylated isoflavones represented by the following formula (1) for producing a nodule formation promoter of chickpea.
  • ⁇ 12> Use of methylated isoflavones represented by the following formula (1) for producing an agent for increasing the yield of chickpea.
  • ⁇ 13> Use of methylated isoflavones represented by the following formula (1) and rhizobia using chickpea as a host for producing a composition for promoting nodule formation or increasing yield of chickpea.
  • the methylated isoflavones are prtensein, biochanin A or formononetin, preferably prtensein.
  • chickpea is preferably a fog species.
  • the agent of ⁇ 16> ⁇ 1>, ⁇ 2>, ⁇ 11> or ⁇ 12> is a composition
  • the content of methylated isoflavones in the composition is preferably the total mass of the composition. 0.000001% by mass or more, more preferably 0.00001% by mass or more, still more preferably 0.0001% by mass or more, and preferably 1% by mass or less, preferably 0.1% by mass or less, more preferably.
  • the number of bacteria rhizobia per composition 1g, preferably 10 4 cfu or more, more preferably 10 5 cfu or more, more preferably 10 6 cfu or more, preferably 10 11 cfu or less, more preferably 10 10 cfu or less, further preferably 10 9 cfu or less, and preferably 5 ⁇ 10 8 cfu or less, preferably or 10 4 to 10 11 cfu, more preferably 10 5 to It is 10 10 cfu, more preferably 10 6 to 10 9 cfu, and preferably 2 ⁇ 10 6 to 5 ⁇ 10 8 cfu.
  • the content of rhizobia and methylated isoflavones is preferably 10 4 to 10 11 cfu and 0.01 to 0.01 to methylated isoflavones per 1 g of the composition.
  • the content ratio of rhizobia and methylated isoflavones is preferably 10 5 ⁇ 5 ⁇ 10 9, more preferably 10 6 ⁇ 5 ⁇ 10 9, more preferably 2 ⁇ 10 6 ⁇ 5 ⁇ 10 9, even more preferably from 2 ⁇ 10 6 ⁇ 5 ⁇ 10 8.
  • the amount of the methylated isoflavones used is preferably 0.000001 to 1 mass ppm as the concentration in the cultivation base material for cultivating chicks.
  • the number of rhizobia to be applied is preferably 10 4 cfu or more, more preferably 10 5 cfu or more, and further preferably 10 6 cfu per chickpea seed. above, and it is preferably 10 7 cfu or more and preferably 10 11 cfu or less, more preferably 10 10 cfu or less, more preferably 10 9 or less, and preferably not more 5 ⁇ 10 8 cfu or less.
  • the amount of methylated isoflavones to be applied is preferably 0.001 ⁇ g or more, more preferably 0.01 ⁇ g or more, and further preferably 0. 1 ⁇ g or more and preferably 1,000 ⁇ g or less, more preferably 100 ⁇ g or less, further preferably 10 ⁇ g or less, and preferably 5 ⁇ g or less, or preferably 0.001 to 1,000 ⁇ g, more preferably 0.
  • the application rate of rhizobia and methylated isoflavones per seed of chicks is preferably 10 4 to 10 11 cfu for rhizobia and methylated isoflavones.
  • rhizobia 10 5 to 10 10 cfu and methylated isoflavones 0.01 to 100 ⁇ g
  • rhizobia 10 6 to 10 9 cfu and methylated isoflavones Classes are 0.1 to 10 ⁇ g
  • preferably rhizobia are 2 ⁇ 10 6 to 5 ⁇ 10 8 cfu and methylated isoflavones are 0.1 to 5 ⁇ g.
  • the ratio of Rhizobium and methylated isoflavones applying is preferably from 10 5 ⁇ 5 ⁇ 10 9, more It is preferably 10 6 to 5 ⁇ 10 9 , more preferably 2 ⁇ 10 6 to 5 ⁇ 10 9 , and preferably 2 ⁇ 10 6 to 5 ⁇ 10 8 .
  • R 1 and R 2 may be the same or different and represent a hydrogen atom or a hydroxy group.
  • Example 1 Effect of methylated isoflavones on root nodule formation and initial growth of chickpeas Early fertilizing soil (Takiisel soil TM-1, Takii Seed Co., Ltd.) and vermiculite fine granules (Akagi Gardening Co., Ltd.) in volume ratio 1
  • the mixture was mixed at a ratio of 1, and about 1.1 L of the soil was filled in a pot (Longpla 120, Nippon Poly Pot Sales Co., Ltd.).
  • Two chickpea (fog seeds) seeds were sown in the pot at a depth of about 1 to 2 cm from the soil surface.
  • Yeast-Mannitol (YM) medium K 2 HPO 4 0.5g, MgSO 4 ⁇ 7H 2 O 0.2g, NaCl 0.1g, Yeast Extract 0.4g, Mannitol 10g, distilled water 1L (pH 6.8)
  • agar Woodo Junyaku Kogyo Co., Ltd.
  • yeast strain NBRC100389T was allowed to stand at 30 ° C. for about 72 hours on the solid medium. It was cultured.
  • the grown rhizobia were inoculated into 50 mL of YM liquid medium in a Sakaguchi flask having a volume of 500 mL, and cultured at 30 ° C.
  • Turbidity OD 600 Rhizobium culture 1mL of the value adjusted with sterile water to 0.02 (7.0 ⁇ 10 6 CFU / ml) of, at the time of seeding, was added dropwise inoculate seeds with micropipettor , Covered with soil.
  • methylated isoflavone and comparative compound were applied as follows. That is, naringenin (Sigma-Aldrich, Cat. No. N5893) and formononetin (Sigma-Aldrich) dissolved in a 0.1 (v / v)% dimethyl sulfoxide (DMSO) aqueous solution so as to be 1 ppm (mg / L). , Cat. No. 47752-5MG-F), Biochanin A (Chromadex, Cat. No. ASB-000002276-005) and Pratensein (Chromadex, ASB-00016080-001) with a micropipettor.
  • DMSO dimethyl sulfoxide
  • a test group to which only 1 mL of 0.1% DMSO aqueous solution was applied was designated as a control group.
  • Cultivation was carried out indoors, and the cultivation conditions were 16 hours in the light period, 25 ° C., an LED light source (Ogetsu Co., Ltd .; model number: VGL-1200W), and a light intensity of 400 to 440 ⁇ mol / m 2 / s. Seven days after sowing, thinning was performed so that there was one plant per pot.
  • Watering was performed by adding a new amount of water to the bat so that the lower part of the pot could be immersed about 5 cm after the bat placed under the pot had run out of water. After cultivating for 25 days, the plant was taken out from the pot, the roots were washed with water, and the nodules were collected. For the collected nodules, the fresh nodule weight and the number of nodules per individual plant were measured. Further, after the plant was dried at 100 ° C. for 3 days, the above-ground dry weight and the underground dry weight of the plant were measured.
  • FIGS. 1 to 4 The measurement results are shown in FIGS. 1 to 4.
  • the graph in each figure shows the mean ⁇ standard deviation. Cont.
  • Nar1, For1, Bio1, and Pra1 represent the control group and the application group of 1 ppm solution of naringenin, formononetin, biochanin A, and prtensein, respectively.
  • the nodule weight tended to increase by 55% in the formononetin group, 26% in the biochanin A group, and 71% in the prtensein group with respect to the control group (Fig. 1).
  • the number of nodules showed an increasing tendency of 15% in the formononetin group, 25% in the biochanin A group, and 38% in the prtensein group, but not in the naringenin group (Fig. 2).
  • the above-ground weight showed an increasing tendency of 19% in the formononetin group, 5% in the biochanin A group, and 23% in the prtensein group with respect to the control group (Fig. 3).
  • Underground weight increased by 34% in the formononetin area, 5% in the biochanin A area, and 11% in the prtensein area, but not in the naringenin area (Fig.). 4).
  • Example 2 Effect of methylated isoflavone on the yield of chickpeas
  • Medium-term fertilizing soil Teakii hydrous cell soil, Takii Seed Co., Ltd.
  • Vermiculite Akagi Gardening Co., Ltd.
  • Example 1 the bacterial solution of chickpea rhizobia (Mesorhizobium ciceri) NBRC100389T strain value was adjusted with sterile water to 0.02 (7.0 ⁇ 10 6 CFU / ml) of the turbidity (OD600) to the seed Inoculated by dropping.
  • Cultivation was carried out from November 2018 to May 2019 in a glass house where sunlight enters.
  • the air conditioning in the house was set to 20 ° C.
  • Eleven days after sowing, thinning was performed so that there would be one plant per pot and that there would be no apparent problem in growth.
  • watering was carried out by irrigating an appropriate amount with a shower once a week, and when the soil surface was found to be dry, additional irrigation was carried out as appropriate.
  • the pods had dried naturally, they were harvested and the grains (beans) were taken out from the pods. For the obtained grains, the dry weight of the grains and the number of grains per individual plant were measured.
  • the measurement results are shown in FIGS. 5 and 6.
  • the graph in each figure shows the mean ⁇ standard deviation.
  • the abbreviations in each figure are the same as those in the first embodiment.
  • the grain weight tended to increase by 5% in the prtensein 0.1 ppm group and 25% in the 1 ppm group with respect to the control group (Fig. 5).
  • the grain number showed a significant increase of 92% in the 0.1 ppm group of prtensein and 55% in the 1 ppm group of prtensein with respect to the control group (Fig. 6).
  • the significance test was performed using the Williams method, and when a significant difference (P ⁇ 0.025) was observed, it was indicated by an asterisk (*).
  • Example 3 shows the results of measuring the protein content contained in the grain of chickpea harvested in Example 2 and converting it into the protein yield.
  • the protein yield tended to increase in the 1 ppm group of prtensein.
  • the protein yield (g / strain) was determined by grain weight (g / strain) ⁇ ⁇ grain protein content (g / 100 g) / 100 ⁇ .
  • the protein content in chickpea grains was measured at the Japan Food Research Laboratories.
  • the combustion method was used for the measurement, and the protein content was calculated by multiplying the quantified total nitrogen by 6.25 as a nitrogen-protein conversion coefficient.
  • Example 4 Synergistic effect of combined use of rhizobia and prtensein Various microorganisms are present in the actual chickpea cultivation soil, and even under such conditions, the effects of rhizobia inoculation and methylated isoflavone are exhibited. Is desired.
  • the combined use of rhizobial inoculation and methylated isoflavone gives root nodule formation under the condition that the soil bacterial solution described below, which is obtained by extracting microorganisms from the field soil, is mixed with the soil, that is, under the condition close to the actual cultivated soil. The impact was assessed.
  • the evaluated test plots 1 to 6 are as follows. Test plot 1. Test plot without soil bacterial solution 2. Test plot with soil bacterial solution 3.
  • the filtrate filtered through 5A was used as a soil bacterial solution, and 45 mL of this bacterial solution was supplied per pot (test groups 2 to 6).
  • Test Group 1 without soil bacterial solution only sterilized water was supplied in the same amount.
  • 200 mL of tap water was separately supplied per pot in order to adjust the water content of the soil.
  • Two chickpea (fog seeds) seeds were sown in each pot at a depth of about 1 to 2 cm from the soil surface.
  • Rhizobium Yeast-Mannitol (YM) medium K 2 HPO 4 0.5g, MgSO 4 ⁇ 7H 2 O 0.2g, NaCl 0.1g, Yeast Extract 0.4g, Mannitol 10g, distilled water 1L (PH 6.8)
  • YM Rhizobium Yeast-Mannitol
  • the grown rhizobia were inoculated into 50 mL of YM liquid medium in a Sakaguchi flask having a volume of 500 mL, and cultured at 30 ° C. with shaking for about 72 hours.
  • the root nodule bacteria culture 1mL of the value adjusted with sterile water to 0.1 (3.5 ⁇ 10 7 CFU / ml) of the turbidity on the day before inoculation (OD 600), at the time of seeding, dropwise to the seed with micropipettor Inoculated (test plots 3 and 4).
  • Results The measurement results are shown in FIGS. 7 to 8.
  • the graph in the figure shows the mean ⁇ standard deviation.
  • the significance test was performed using the Tukey-Kramer method, and different letters (a, ab, b) between the groups indicate that a significant difference (P ⁇ 0.05) was observed. Almost no nodules were formed in the test groups 1 and 2 and the test group 6 in which the prtensein was applied alone. In Test Group 3, the number of colonies increased by inoculation with rhizobia, but the effect was limited.
  • the number of nodules is set to the test group 3 respectively.
  • the nodule weights were 2.3 times and 2.2 times in the test groups 4 and 5, respectively, with respect to the test group 3 (FIG. 8).

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Abstract

La présente invention concerne une technique consistant à favoriser la formation de nodules par des bactéries de nodules qui coexistent avec des pois chiches et l'accroissement du rendement des pois chiches.　Le promoteur de formation de nodules de pois chiche contient, comme ingrédient actif, des isoflavones méthylés représentés par la formule (1) (dans la formule, R1 et R2 peuvent être identiques ou différents, indiquant chacun un atome d'hydrogène ou un groupe hydroxy).
PCT/JP2020/044924 2019-12-02 2020-12-02 Promoteur de formation de nodules de pois chiche Ceased WO2021112139A1 (fr)

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CN116694496B (zh) * 2022-12-07 2024-09-13 塔里木大学 一种促进鹰嘴豆结瘤的奇台根瘤菌及其用途

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CN115644351B (zh) * 2022-11-14 2024-05-14 湖北工业大学 提高工业酵母发酵活性的方法

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