JP4490696B2 - Degradation accelerator and method for promoting degradation of coated gel layer of gel-coated seed - Google Patents

Description

  The present invention relates to a decomposition accelerator for a coated gel layer of a gel-coated seed.

Gel-coated seeds consist of plant seeds and seed analogs such as viable embryos or somatic embryos (hereinafter referred to as "seed"), water as a dispersion medium, and water and dispersoids. It is encapsulated in a coated gel layer made of an aqueous gel, and it is an epoch-making stage that can reliably obtain seedlings from a single seed and eventually harvest by the moisture and other components constituting the coated gel layer. Because of this certainty, it is said that it is possible to use an excellent but extremely expensive F ₁ seed, and as a result, a significant improvement in yield and quality can be achieved.

  Such a gel-coated seed is, for example, an aqueous solution containing metal ions that solidify the gel-forming aqueous solution by encapsulating the seeds in a gel-forming aqueous solution containing sodium alginate as a main component, for example, a divalent or higher-valent metal ion. For example, it is made by gelling by reacting sodium alginate around the seeds with these metal ions by dropping them into an aqueous solution of a salt of calcium, barium, aluminum or the like.

  Here, the coated gel layer of the gel-coated seed is required not to be crushed in handling until sowing and to be excellent in handling, but after sowing, a property of quickly losing elasticity and decomposing is required. .

  In practice, however, the coated gel layer that wraps the seeds by drying after sowing loses moisture and changes to a hard film after drying, which inhibits the growth of shoots and roots from the seeds, depending on the type of plant. Causes problems such as inferior seedling compared to normal seeds. In order to prevent such obstacles, gel-coated seeds require more attention to irrigation after sowing than normal seeds, and there is a problem that labor is required in actual cultivation.

  In order to solve this problem, there has been proposed a method of chemically disintegrating the coated gel layer by using a drug (chelating agent) having a chelating ability with respect to calcium ions reacting with alginic acid. At this time, the chelating agent affects the electrical conductivity in the soil, causing problems such as poor plant seedling establishment, and it is judged that the chelating agent is inappropriate as a solubilizer for gel-coated seeds.

In addition, the present inventors have proposed a technique that can disintegrate a coated gel layer very effectively even with a small amount of a chelating agent by a microcapsule having thermoreversibility in JP-A-9-15. However, in this case, the occurrence of plant seedling failure can be suppressed to a very low level, but since heat treatment is required, there is a problem that it cannot be applied to heat-sensitive plant seeds.
Japanese Patent Laid-Open No. 9-15

  The present invention improves the above-mentioned conventional problems, that is, does not require heat treatment, and quickly decomposes the coated gel layer of the gel-coated seed without causing problems such as plant seedling failure. An object is to provide a method capable of

In order to solve the above-mentioned problem, the agent for promoting the degradation of a coated gel layer of a gel-coated seed according to the present invention comprises a gel-coated seed comprising a coated gel layer made of an aqueous gel having an alginate as a dispersoid as described in claim 1. A coating of a gel-coated seed characterized by containing Pseudoarteromonas Atlantica as a microorganism that produces an enzyme that decomposes the dispersoid that constitutes the aqueous gel of the coated gel layer. It is a gel layer decomposition accelerator.

Accelerated degradation process of gel coat of gel-coated seeds of the present invention is to solve the above problems, as described in claim 2, promote decomposition of the gel coat of gel-coated seeds having a gel coat comprising an aqueous gel A method for promoting the degradation of a coated gel layer of a gel-coated seed, wherein the degradation-promoting treatment is carried out by an enzyme that degrades the dispersoid constituting the aqueous gel of the coated gel layer, wherein the gel-coated seed constitutes the coated gel layer The alginate has an alginate as a dispersoid of the aqueous gel, and the enzyme that degrades the dispersoid is an alginate-degrading enzyme, and a liquid containing the alginate-degrading enzyme is sprayed on the gel-coated seed after sowing. A method for promoting the degradation of a coated gel layer of a gel-coated seed, wherein the degradation promoting treatment is performed .
The method for promoting the degradation of the coated gel layer of the gel-coated seed according to the present invention is the method for promoting the degradation of the coated gel layer of the gel-coated seed provided with the coated gel layer made of an aqueous gel. A method for promoting the degradation of a coated gel layer of a gel-coated seed, comprising contacting Pseudoalteromonas atlantica as a microorganism that produces an enzyme that degrades a dispersoid that constitutes an aqueous gel of the gel layer. is there.
The method for promoting the degradation of a coated gel layer of a gel-coated seed according to the present invention is the method for promoting the degradation of a coated gel layer of a gel-coated seed provided with a coated gel layer comprising an aqueous gel. A method of accelerating the degradation of a coated gel layer of a gel-coated seed, which is subjected to a degradation promoting treatment by an enzyme that decomposes a dispersoid constituting the aqueous gel of the gel layer, wherein the gel-coated seed disperses the aqueous gel that constitutes the coated gel layer The above-mentioned enzyme that decomposes the dispersoid is alginic acid-degrading enzyme having an alginate as a quality, and an aqueous solution having a concentration of the enzyme of 0.1 U to 4 U / mL is used before seeding or at the time of sowing. This is a method for promoting the degradation of a coated gel layer of a gel-coated seed, characterized by performing a degradation promoting treatment on the gel.

  According to the agent for promoting the degradation of the coated gel layer of the gel-coated seed according to the present invention, the coated gel layer of the gel-coated seed can be rapidly decomposed without causing problems such as plant seedling failure.

  Generally, a gel is composed of a dispersoid that forms a molecular network to maintain its shape, and a dispersion medium whose degree of freedom is significantly limited by the dispersoid (in the case of an aqueous gel, the dispersion medium is water). However, the enzyme used in the present invention must be capable of degrading this dispersoid.

  Here, what can be used as the dispersing agent of the aqueous gel constituting the coated gel layer of the gel-coated seed is, for example, duran gum, xanthan gum, carboxymethyl cellulose (CMC), pectin, gelatin, locust bean gum, carrageenan, polyacrylic. Examples include sodium acid and agar.

  Here, as the enzyme used in the present invention, in addition to the property capable of decomposing the dispersoid, it is necessary not to inhibit seedling establishment and to have no adverse effect on the environment. It is preferable that it is water-soluble because contact is facilitated.

  In the present invention, the decomposition of the dispersoid is to break the aqueous gel formed by the dispersoid and the dispersion medium by breaking the molecular chain constituting the dispersoid, and this decomposition causes the aqueous gel of the coated gel layer to flow. Although it is most desirable to be a sol having properties, it should be soft enough to prevent the growth of plants in the coated gel layer.

  Here, for example, among combinations of a dispersoid that constitutes an aqueous gel together with water and its degrading enzyme, combinations that can be used in the field of gel-coated seeds include alginic acid and its salts and alginic acid degrading enzyme, carboxymethylcellulose and cellulase, agar And combinations of agarase and the like. In addition, although these combinations may be used independently, as long as the effect of this invention is acquired and it does not exert a bad influence on the plant for cultivation purpose, you may combine multiple sets.

  Among these, in the combination of alginic acid and its salt as a dispersoid constituting the aqueous gel of the coated gel layer and an alginate degrading enzyme as an enzyme that degrades the dispersoid, an alginate molecular chain (as the dispersoid constituting the aqueous gel) In the case of a coated gel layer having a molecular chain of alginic acid or a salt thereof, alginic acid lyase is used as the alginate decomposing enzyme, and the alginic acid molecule is decomposed by the degradation accelerator of the coated gel layer of the gel-coated seed containing the alginate lyase. The aqueous gel layer forming the coated gel layer can be disrupted by lowering the molecular weight by oligomerization or monomerization. This alginate lyase does not adversely affect plant seedling establishment.

  In addition, in the combination of alginic acid and its salt as a dispersoid constituting the aqueous gel of the coated gel layer and an alginate-degrading enzyme as an enzyme that decomposes the dispersoid, by research of Matsubara et al. According to the configuration of the present invention, fertilization is performed by decomposing the coated gel layer, as reported that alginic acid oligosaccharide obtained as a degradation product of alginic acid by alginic acid-degrading enzyme promotes plant root elongation. Equivalent effects can be obtained.

  In the present invention, the enzyme that degrades the dispersoid is, for example, a solution prepared by dissolving the enzyme in an appropriate solvent (preferably water) from the outside of the coated gel layer of the gel-coated seed to the coated gel layer. At this time, it may be enclosed in a microcapsule or the like made of a material that is gradually soluble in the solvent, and may be present in advance when the coated gel layer is formed. In the case of a gel-coated seed of a seed having a relatively high heat resistance, a coated gel layer using a microcapsule using a thermoreversible aqueous gel (a substance enclosed inside is released when heated). It is also possible to use means such as heating the coated gel layer at the start of the acceleration of decomposition. Further, a means such as injecting an enzyme solution into the coated gel layer of the gel-coated seed using a hollow needle or the like can also be used.

  In the case where a solution in which such an enzyme is dissolved is used as an agent for promoting the degradation of the coated gel layer of the gel-coated seed, the method of contacting the coated gel layer from the outside of the coated gel layer of the gel-coated seed is contacted. It can be divided into three according to the time.

  A pre-seeding contact treatment method in which a solution in which an enzyme is dissolved before sowing is contacted with a gel-coated seed provided with a coated gel layer by normal means such as immersion, spraying, and spraying, and a decomposition promoting treatment is performed, When sowing, contact method at the time of sowing in which the solution in which the enzyme is dissolved by the same means is brought into contact with the gel-coated seed, or a solution in which the enzyme is dissolved in the gel-coated seed after sowing to the cell tray or the field. There are three post-seeding contact treatment methods that perform spraying and promoting decomposition.

  In the above pre-seeding contact treatment method, when the gel-coated seeds are immersed in a solution in which the enzyme is dissolved before sowing, the longer the time of immersion, the more the degradation of the coated gel layer proceeds, and the faster the decomposition after sowing. However, as the decomposition progresses, the handling at the time of sowing deteriorates, or obstacles such as the inability to machine sowing occur. Seeding while the progress of disintegration of the gel layer can be sowed.

  In addition, when the solution in which the enzyme is dissolved is brought into contact with the gel-coated seed by spraying, spraying, or the like in the contact treatment method before seeding, the solution in which the enzyme is dissolved can be selected by selecting conditions such as the spraying amount and the solution concentration. In general, the coated gel layer can be kept in a state where it can be mechanically seeded within a few hours after contact with the liquid, but the handling time at the time of sowing deteriorates as long as the time from contact with the liquid to sowing is long. Therefore, it still needs to be examined in advance.

  In the contact treatment method at the time of sowing, since the solution in which the enzyme is dissolved is brought into contact with the gel-coated seed at the time of sowing by means such as spraying, spraying, and dipping, the above-mentioned problems in the contact treatment method before sowing usually do not occur. .

  On the other hand, in the contact treatment method after sowing, there is a method of spraying a solution in which an enzyme is dissolved in a gel-coated seed after sowing in a cell tray or a field. In this lifting, the handling property of the gel-coated seed is maintained with respect to the sowing operation, but more solution in which the enzyme is dissolved is required as compared with the above two methods.

  In addition, in the contact treatment method after sowing, when the soil seeded with the gel-coated seed is dry, the solution in which the sprayed enzyme is dissolved is absorbed into the soil, and as a result, the decomposition effect on the gel-planted seed is reduced. There is a possibility that after sowing, irrigate and spray the solution in which the enzyme is dissolved, or sow the irrigated soil and spray the solution in which the enzyme is dissolved before the soil dries. Thus, it is desirable that the soil contains a certain amount of water when the solution in which the enzyme is dissolved is sprayed.

  Whether or not to cover the seeded gel-coated seed is appropriately determined depending on the nature of the seed plant enclosed in the gel-coated seed.

  In addition, the present invention does not include an enzyme directly in the degradation accelerator for the coated gel layer of the gel-coated seed of the present invention, but contains a microorganism that produces an enzyme that degrades the dispersoid constituting the aqueous gel of the coated gel layer, You may produce the enzyme which decomposes | disassembles the dispersoid which comprises the aqueous gel of a coating gel layer with this microorganism.

  As long as the effect of the present invention is obtained, the gel-coated seed used in the present invention is a substance that forms other gel-like substances, for example, various synthetic and semi-synthetic water-absorbing polymers (starch, polyacrylate, cellulose, etc.) , Or gel-like dispersed particles (particles of a size that is not normally visible) made of a natural gel-forming polymer such as agar and water, or other components that contribute to the growth of the plant seeds contained For example, a fertilizer component (which may be solid or may be dissolved in water in the coated gel layer as a component), or air or oxygen may be disposed in the coated gel layer.

Examples of the decomposition accelerator for the coated gel layer of the gel-coated seed of the present invention will be specifically described below.
«Examples in combination using alginic acid and its salt as a dispersoid constituting the aqueous gel of the coated gel layer and alginate lyase as an enzyme that degrades these dispersoids»
First, an example of a combination using alginic acid and its salt as a dispersoid constituting the aqueous gel of the coated gel layer and alginate lyase as an enzyme for decomposing these dispersoids will be described.

  In this example, an alginate lyase aqueous solution was used as an agent for promoting the degradation of the coated gel layer of the gel-coated seed. However, in preparing these alginate lyase aqueous solutions, it is necessary to pay attention to the following points. That is, since commercially available alginate lyase has different enzyme activity for each product, a desired concentration cannot be exhibited at a weight percent concentration when setting a concentration having effective activity. Therefore, in the present invention, the enzyme amount concentration (U / mL) contained in the alginate lyase aqueous solution per mL was used as the concentration unit.

  Here, the unit “U (unit)” of the enzyme amount is defined as follows. That is, 0.15 mL of enzyme solution was added to 4.5 mL of 0.1% alginate solution (0.2 M phosphate buffer, pH 6,3), reacted at 37 ° C. for 30 minutes, and then 4.65 mL of 0.1 M NaOH. The amount of enzyme that increases the absorbance at a wavelength of 235 nm per mL of the reaction solution by 1 minute is defined as 1 U.

  In the combination using alginic acid and a salt thereof as a dispersoid constituting the aqueous gel of the coated gel layer and alginate lyase as an enzyme that degrades these dispersoids, the concentration of the aqueous alginate lyase solution used is 0.1 U / mL or more. It is desirable to be. If the concentration is less than 0.1 U / mL, the degradation effect of the gel having alginate is hardly obtained, and as a result, the effect of increasing the seedling establishment rate is small. In addition, the higher the enzyme concentration in the aqueous solution, the higher the degradation effect. However, in the range of 0.1 U to 4 U / mL, no significant difference was observed in the final seedling establishment rate. We cannot expect improvement in standing. And at this time, since the usage-amount of alginate lyase increases and it becomes high-cost, it is desirable that the upper limit of the density | concentration of the alginate lyase aqueous solution to be used is 4 U / mL or less.

  As a guideline, the amount of alginate lyase aqueous solution applied to the gel-coated seeds after sowing to the field or cell tray is 1 mL of the alginate lyase aqueous solution in the above concentration range per gel-coated seed. However, since the amount of liquid greatly affects the workability at the time of spraying rather than the effect on the acceleration of gel degradation, the minimum necessary amount should be examined in advance by considering the presence or absence of soil covering after sowing. Determine the actual application amount while taking into consideration.

  In addition, when using an alginate lyase aqueous solution with a concentration of 1 U / mL, the number of spraying is usually sufficient just after seeding, but when dividing the spraying into multiple times, the gel is dried to dry the lyase. Since the degradation effect is reduced, it is desirable to finish the treatment within 2 days after sowing. The relationship between the concentration and the number of times is examined in advance.

In this example, a microorganism (such as Pseudoalteromonas atlantica) that produces alginate lyase instead of the alginate lyase is brought into contact with the coated gel layer as a decomposition accelerator for the coated gel layer of the gel-coated seed. This case may also be included in the present invention. At this time, it is desirable to contact a medium or / and a culture solution containing an essential component for culturing the microorganism together with these microorganisms, or use a gel-coated seed containing such an essential component in a coated gel layer.
<Experimental conditions>
Calceolaria gel-coated seeds Used seeds: Calceoraria F1 Midas Temperature conditions: constant 20 ° C, 12 hours day covering Soil after sowing: None <Production of gel-coated seeds>
Weigh out sodium alginate and powdery starch / polyacrylate water-absorbing polymer so that the concentration in the solution is 0.9% by weight and 0.2% by weight, respectively, add water and stir well to form gel. A liquid was prepared. When the water-absorbing water-absorbing polymer particles dispersed in the gel-forming liquid in which the particles comprising the water-absorbing water-absorbing polymer in the liquid were dispersed were observed with a microscope, the diameter was 0.2 mm or less and 0.1 mm or more, and the average was It was 0.15 mm.

Using this gel-forming liquid, the droplet is formed at the lower end of the hollow tube, and seeds are introduced into the droplet by supplying calceolaria seeds into the droplet from the inside of the hollow tube. Were dripped into an aqueous calcium chloride solution (cured solution) having a concentration of 10% by weight, respectively, to obtain a gel-coated seed obtained by arranging seeds in a coated gel layer composed of an alginate-based aqueous gel. In addition, the size of these obtained gel-coated seeds is a substantially spherical shape having a diameter of about 1 cm. In addition, the addition of a water-absorbing polymer to the coated gel layer as described above gives good seedling establishment while having sufficient strength for handling both immediately after production of the gel-coated seed and after restoration after storage drying. It is to do.
<Implementation procedure>
The gel-coated seeds prepared above are sown in a cell tray (288 holes, the soil is Metromix 350 (manufactured by Scotts)). Next, irrigation is carried out to wet the periphery of the gel-coated seed.

Immediately after that, an aqueous solution of alginate lyase prepared to a concentration of 0.4, 1, 2, or 4 U / mL is sprayed uniformly on the cell tray with 1 mL per gel-coated particle. In addition, it sprayed with respect to 48 gel-coated seeds using the density | concentration of each alginate lyase aqueous solution. After spraying, cultivation was carried out as usual.
<Evaluation>
Results of using alginate lyase aqueous solution prepared at a concentration of 0.4, 1, 2, or 4 U / mL, which is a degradation accelerator for the coated gel layer of the gel-coated seed according to the present invention, regarding the transition of seedling establishment after sowing (Results of gel-coated seeds that were seeded at the same time (“0.4 U / mL”, “1 U / mL”, “2 U / mL”, or “4 U / mL”, respectively) without spraying an alginate lyase aqueous solution) It is shown in FIG.

  According to FIG. 1, when the decomposition accelerator for the coated gel layer of the gel-coated seed according to the present invention is used, an alginate lyase aqueous solution having a concentration of 0.4, 1, 2, or 4 U / mL is used. Even if it exists, it turns out that seedling establishment is remarkably improved compared with the case where the process by the decomposition | disassembly promoter of the coating gel layer of a gel coating seed is not performed.

  In addition, in the above experimental process, when the degradation accelerator for the coated gel layer of the gel-coated seed according to the present invention was used, it was observed that the degradation of the coated gel layer of the gel-coated seed was good. When the decomposition accelerator for the coated gel layer of the coated seed was not used, it was observed that the coated gel layer did not decompose and covered the seed as a hard film.

  According to the present invention, in the gel-coated seed, the coated gel layer of the gel-coated seed can be quickly decomposed without causing problems such as plant seedling failure.

The results of examining the history of seedling establishment rate after sowing when using the coating accelerator for the gel-coated seeds of the gel-coated seed according to the present invention, when the decomposition accelerator for the gel-coated seed coating gel layer is not used It is the graph shown with the result.

Claims (4)

Microorganism that produces an enzyme that degrades the dispersoid that constitutes the aqueous gel of the coated gel layer in the degradation accelerator for the coated gel layer of the gel-coated seed comprising a coated gel layer comprising an aqueous gel having an alginate as a dispersoid A degradation accelerator for a coated gel layer of a gel-coated seed characterized by containing Pseudoarteromonas Atlantica as In a method for promoting the degradation of a coated gel layer of a gel-coated seed comprising a coated gel layer made of an aqueous gel, the coating of the gel-coated seed is subjected to a degradation promoting treatment with an enzyme that degrades a dispersoid constituting the aqueous gel of the coated gel layer. A method for promoting the degradation of a gel layer,
The gel-coated seed has an alginate as a dispersoid of the aqueous gel constituting the coated gel layer, the enzyme that degrades the dispersoid is an alginate-degrading enzyme, and
A method for promoting the degradation of a coated gel layer of gel-coated seeds, wherein the degradation-promoting treatment is performed by spraying a solution containing the alginate-degrading enzyme on the gel-coated seeds after sowing . In a method for promoting the degradation of a coated gel layer of a gel-coated seed comprising a coated gel layer made of an aqueous gel, Pseudoarteromonas Atlantica is used as a microorganism that produces an enzyme that degrades the dispersoid constituting the aqueous gel of the coated gel layer. A method for promoting the degradation of a coated gel layer of a gel-coated seed, comprising bringing the coated gel layer into contact with the coated gel layer. In a method for promoting the degradation of a coated gel layer of a gel-coated seed comprising a coated gel layer made of an aqueous gel, the coating of the gel-coated seed is subjected to a degradation promoting treatment with an enzyme that degrades a dispersoid constituting the aqueous gel of the coated gel layer. A method for promoting the degradation of a gel layer ,
The gel-coated seed has an alginate as a dispersoid of the aqueous gel constituting the coated gel layer, and
The enzyme that degrades the dispersoid is an alginate-degrading enzyme, and an aqueous solution having a concentration of the enzyme of 0.1 U to 4 U / mL is used to perform degradation promoting treatment on the gel-coated seed before or during sowing. A method for promoting the degradation of a coated gel layer of a gel-coated seed, which is characterized .

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