JPS60180521A - Culture of mushrooms - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention involves culturing the mycelium of mushrooms, that is, basidiomycetes, by a dialysis culture method, and using the mycelium obtained by the method to accelerate the growth of fruiting bodies and to reduce the total production. Concerning methods of cultivating mushrooms to improve

In the present invention, target mushrooms used for cultivation include shiitake, oyster mushroom, namekotake, enokitake, wood ear mushroom,
Any mycelium of basidiomycetes such as Argentinian fungus, Stonewort mushroom, Maitake mushroom, Pine mushroom, Fukurotake mushroom, Grouper mushroom, Shimeji mushroom, etc. may be used.

Thirteen fruiting bodies of the above mushrooms are currently being cultivated artificially, and many of them are relatively easy to obtain. and,
Including the cultivation period, it takes about 340 days to harvest these fruiting bodies in the case of cutlet cultivation and about 280 days in the case of bottle cultivation.

However, there is still no known method for harvesting the above-mentioned fruiting bodies in a short period of time with a high yield.

The present invention proposes an efficient method for producing fruit bodies, which has not been possible using conventional methods. Incidentally, according to the present invention, the period in the fruiting body harvesting box is 18
It is 0 days.

Traditionally, the most common method for cultivating basidiomycetes is to mix sawdust, rice bran, lignin, agar, etc. with water to create a medium, fill the mixed medium in bottles, and after sterilization, aseptically incubate the mother. This is a method in which the fungus is implanted in a culture medium to create mycelia that serve as a seed fungus, and the mycelia are allowed to germinate in a bottle to cultivate fruiting bodies.

In this case, the optimum humidity and moisture content of the culture medium is such that when lightly squeezed by hand, some water drips out, and this is the practice.

However, the optimal culture medium humidity and moisture conditions in this case are only those at the time of preparing the culture medium, and the humidity and water content in the culture medium will change rapidly due to the subsequent heating and cooling operations required for sterilization, and the hyphae will grow. It is difficult to maintain optimal prefectural boundaries for growth.

Therefore, these conditions after sterilization are deficient in moisture, which inhibits the short-term spread of mushroom mycelia, which is also bad for the growth of mushroom fruiting bodies, and reduces the yield of mycelia and fruiting bodies. [7] The disadvantage is that you cannot expect the best.

It is well known that when cultivating mushrooms, if hyphae are allowed to spread in a medium over a short period of time, hyphae with high mushrooming activity will be obtained and the production of fruiting bodies will be high. In order to make hyphae grow in a short period of time, it is necessary to adjust the culture medium conditions, particularly humidity, moisture, and temperature, so that they are optimal for the spread of hyphae.

However, in order to accelerate the growth of fruiting bodies and improve production, it depends on how to accumulate mushroom-starting activity during cocoon filament culture.

As a result of extensive research to remove the conventional drawbacks and make use of the above knowledge, the present inventor has discovered that the humidity and moisture in the culture medium,
It is said that mycelium obtained by the dialysis culture method, which causes temperature changes over time and is easy to sterilize, has a significantly higher yield of fruiting bodies than mycelia obtained by conventional methods. The inventors discovered the effect and arrived at the present invention.

In the present invention, a paste-like medium prepared by adding a small amount of sugar to sawdust, rice bran, potatoes, or corn that has been sterilized in advance by a conventional method is applied to a medium 9-fiber membrane in a sterilized hollow fiber dialysis container. Aseptically insert it into the outer wall side of the

These mediums are applied to the outer wall surface of each inner membrane so as to be approximately uniform, and air is allowed to flow in aseptically. On the other hand, inside the medium 9 system, water necessary for the mycelium is circulated aseptically and the temperature is set in the range of 15 to 40C. This makes it possible to prevent aging of moisture, which could not be solved by conventional methods.

By this method, the humidity in the culture medium is set to an environmental condition in which the relative humidity is 80% or more and the vortex layer is 15 to 40 c, so that mycelium can grow with sufficient activity. Preferred conditions for further increasing the activity are a relative humidity of 90% or more and a temperature in the range of 25 to 35C.

Next, the culture medium adhered to the outer wall surface of the inner fiber by the above method is made to support the tissue-cultured seed bacteria in a sterile manner while allowing air to flow into the culture medium. In this way, under completely sterile conditions, a voltage of 7V is generated between the container inside the dialysis container and the outer wall 11Q of the medium perforation.
The space filled with hyphae becomes a closed space, and the hyphae can be spread over the entire semi-permeable outer wall for a long time. It is this hyphae that exhibits great mushrooming activity and promotes the growth of fruiting bodies in the next cultivation stage.

The object of the present invention is to obtain mycelium with high mushrooming activity through dialysis culture and use it as a seed to cultivate fruiting bodies with high growth rate.

It should be noted that the dialysis type culture container requires an inlet/outlet for introducing and removing the culture medium, various nutrients, and air in order to carry out the culture reaction. To this end, the main case is provided with an inlet/outlet, and through this inlet/outlet, Qi, culture medium, and nutrients are aseptically introduced into and taken out of the outer wall surface of the hollow fibers and the mycelial culture zone inside the main case. After the addition of culture medium and nutrients is completed, this inlet/outlet must be sealed with a plug that does not allow germs to pass through but allows air to pass through so that air can flow into the culture medium side in an aseptic manner.

Furthermore, the average pore diameter of Nakai Ito with a built-in cultured mustard is 600X~
A membrane with a thickness of 2.0 μm, preferably 500^ to 1.5 μm is used, and moisture is introduced from inside the medium 9 system.

In the present invention, the hollow fiber type dialysis container used is composed of a main body case having an inlet/outlet for a medium 9 fiber, 9 qi or nutrients, and an element, and a header camp. The materials constituting each part may be of any material, but the most common ones are cellulose-based hollow fibers for hollow fibers, acrylonitrile styrene copolymer for the case body, and polypropylene for the header cap.

However, compared to the inner cross-sectional area of the main body case containing the hollow fibers, the hollow fiber cross-section (including both the hollow fiber membrane and the inner part)
When the cross-sectional area occupied by the microorganism exceeds 70%, the growth of mycelium seems to slow down. Therefore, the 9 spaces formed by the main body case and the cross section of the hollow fiber are necessary as the 9 spaces in which the hyphae grow.

The hollow fiber membrane in the dialysis culture container may be either a regenerated cellulose-based membrane or a synthetic polymer-based membrane;
In particular, hollow fiber membranes made of Cubula ammonium rayon generate mycelium in a short period of time, and in large quantities. Hollow fiber membranes made of synthetic polymers have less hyphae than cuproammonium rayon membranes. The reason for this is obvious, but it is presumed that the cellulose membrane functions better as a medium or support for the hyphae.

In order to prove the above fact, the following experiment was conducted.

That is, cupraammonium rayon, cellulose-based membrane/acetate, and polyacrylonitrile membranes were selected as membrane materials, and hyphae were cultured using the above-mentioned culture conditions for each membrane.

As a result, a large amount of white mycelium occurs on the outer wall surface of the cupraammonium rayon membrane.

A small amount of hyphae was observed on the cellulose diacetate membrane, but no hyphae were observed on the polyacrylonitrile membrane. Since no hyphae were observed on the so-called synthetic polyacrylonitrile membrane, and hyphae were observed on cuproammonium rayon, we thought that the presence of cellulose might be involved in the hyphae generation. We investigated the coexistence of carriers that have an affinity for hyphae.

The fruiting body residue from which the active ingredient had been extracted was used as a carrier. This fruiting body residue is obtained by purifying and separating the active ingredients of the basidiomycete cultured on the outer wall of the hollow fiber into the inside of the hollow fiber by introducing an extractor into the outer wall of the hollow fiber. However, when this is used as a carrier, hyphae are also observed on cellulose diacetate membranes and polyacrylonitrile membranes under the same culture conditions.

However, the amount of mycelium that develops on the cupra ammonium rayon membrane cultured without fruiting body residue in the culture is not large enough to completely fill the culture container, but the amount of mycelium is comparable to that of kanashi. Hyphae can be cultured.

There is no significant difference in the amount of hyphae generated between cellulose diacetate membranes and polyacrylonitrile membranes, depending on the coexistence of the carrier.

The clump bacteria cultured by the dialysis culture method described in A+J may be cultivated in the same manner as normal hydroponic cultivation.

That is, an old oak or beech tree that has been cut into approximately 15 square meters in diameter and approximately 15 square meters in length is sterilized. The surface of this sterilized old tree is evenly sprinkled with inoculum cultured by dialysis in an aseptic manner. When these scraps covered with mycelium are sealed with a vinyl sheet and stored at 25C±1 for about 50 to 80 days, white mycelium grows on the surface of the scraps.

At this point, both the original and the original are sealed inside the greenhouse using the usual method. During this sedation, a material with good water retention and readability is applied to the surface of the tank water where white mycelia are growing in clusters, and primordia are generated. If this material becomes too thick, it will inhibit the development of fruiting bodies.

The temperature inside the greenhouse is maintained within the range of 60C ± 5C, and after germination of the primordia is observed, water is sprinkled once every other day to maintain relative humidity at 80% or higher. Growing fruit bodies are obtained 100 to 180 days after the start of sedation.

An interesting finding was obtained during this process.

That is, in the dialyzed cultured hyphae according to the present invention, primordia-like masses that are an inhibitory factor for the growth of fruiting bodies do not occur. In cultivation using other methods, two such clumps were generated and did not affect the growth of the fruiting body. Once this primordium-like mass occurs, the growth of fruiting bodies around the primordium stops, and the surrounding fruiting bodies that are still growing are eaten into the primordium mass. It is eaten six times and remains as an immature fruiting body primordium.

Therefore, it prevents the primordium-like mass from breaking down and the normal development of the fruiting body, reducing the productivity of the fruiting body. Naturally, the fact that this primordium mass does not occur in the inoculum group produced by the dialysis culture method leads to an improvement in the productivity of fruiting bodies.

It is not clear why these lumps do not occur in the dialysis culture method, but it is presumed that sterilization during mycelial culture and the concentration of oxygen and water are affected.

In the dialysis culture method according to the present invention, the concentration of oxygen and water is higher than in other methods, and the hyphae grow in a sterile state.
This is because it has been cultivated to a high degree of susceptibility.

Comparative Examples and Examples are shown below to specifically illustrate the effects of the present invention.

Example: Thoroughly boiled potato quoyvtc, 101 grams of corn germ powder, 101 grams of sugar and 200 W of water are added to make a paste-like medium and sterilized.

In this sterilized medium, the original fungus of C. chinensis is cultured, and the mycelium is multiplied. The grown mycelium was used as a seed fungus, and the culture medium was mixed with sterilized sawdust 1501.
0 [aseptically put into a wide-mouthed bottle to grow mycelia.

The growth conditions are a humidity of 80% or higher and a temperature of 2o to 25C. When left for 25 days, clusters of white mycelium can be seen on the walls of this wide-mouthed bottle. This wide mouth bottle sample is %1
shall be.

Using mycelia grown in the same medium as used in the case of Nn1 as a seed fungus, the medium and the medium are uniformly sprinkled on the circular cross-sections of four old beech trees that are sterilized and have a radius of 15 Qyx and have been sufficiently absorbed with sterile water. .

Seal the above building blocks sprinkled with mycelium with a pilar sheet, and
When stored at 5C for 80 days, white mycelium grows on the surface of the tank water. This will be the slow 2 sample.

In addition, the temperature of a steam-sterilized hollow fiber dialysis container (manufactured by Asahi Medical #f) with a surface area of 0.8rn2 was maintained at 25C, sterile hot water was poured inside the hollow fiber, and N was applied to the outer wall of the hollow fiber.
The hyphae grown in the same medium used in n1 are partially implanted on the outer wall surface of the hollow fiber together with the medium. After 6 days, white hyphae grow in clusters over the entire surface of the outer wall of the hollow fibers.

This is designated as sample No. 6.

Mycelium is collected for each case of N[l 1 , Nn 2 , and Noj, dried, and weighed. Table 1 shows the weight (harvest amount) of C. chinensis mycelium per 1 m area at that time.

Table 1: Yield of 4 volumes of Cinnamon mycelia The mycelium obtained in each case shown in Table 1 was collected, and the fruiting bodies were cultivated using the usual Sabuto method as described below.

That is, 324 old beech trees each having a cylindrical shape of 15 meters in diameter and 15 meters in height were sterilized in a high-pressure cooker.

The processing conditions at that time were a temperature of 105 to 110C.

The pressure is 0.4 k7/Crn2. Next, three mycelia shown in Table 1 were collected, and 1 cocoon filament was collected per cocoon thread of each type.
08 trees are prepared, and the mycelia of 5 trees are combined to make a total of 6.
24), and implant mycelium on each one. The implantation surface spans a flat surface with a diameter of 15 mm, and the mycelium is spread over this flat surface, and the hyphae are implanted at a temperature of 25C ± 5C1 with a relative humidity of 80% or more to activate the mycelium and germinate the fruiting body. The cells are left in a culture room maintained under a constant environment for 80 days.

This cultured sagi was placed in a plastic greenhouse maintained at a temperature range of 3oC±50℃, and one primordium germinated.

Once the primordium has germinated, water it every other day and observe the degree of germination and growth of the fruiting body.

After 180 days have elapsed since the start of taming, the growing fruit bodies are harvested and the yield is examined at the same time. The results are shown in Figure 2.

Table 2 Fruiting body development status and yield amount As is clear from Table 2, there are differences in the yield of cultivated fruiting bodies depending on the mycelial culture method used before inoculating the seed fungus into the tank water. The mycelium obtained by the dialysis culture method according to the present invention has the highest yield, followed by the bottle culture, and finally the block culture.

Also, during the above-mentioned sedation period, even when the fruiting bodies begin to appear, the white germinated mycelial mass still remains.
There is a lump that remains as an immature primordium and continues to grow.This is an immature fruiting body that cannot grow into a fruiting body (has no mushrooming activity), but the fruiting body Instead of growing as a mycelium, the surface remains mycelium-like, and it continues to multiply, absorbing nutrients from the fruiting bodies growing around it, and even consuming the fruiting bodies. They are like so-called hyphal cancer cells.

Therefore, when this white mass occurs, the growth of fruiting bodies in the surrounding colonies is inhibited, and as a result,
There is a decrease in the number of rats producing harvested fruiting bodies.

These facts indicate that there are differences in the number of white germinated chisel masses that occur depending on the culture method of the inoculum, and depending on the degree of occurrence of these,
This will affect the yield of fruiting bodies. In this case as well, it can be seen that the dialysis culture method is superior as a mycelial culture method.

Claims (2)

[Claims] (1) On the outer wall surface of the aseptically treated hollow fiber membrane, one solid foot of the basidiomycete Kanakura is aseptically implanted with a paste-like medium, and at the same time, air is circulated and inflowed in an aseptic manner, while the hollow fiber membrane is Sterilized water is circulated inside and brought into contact with basidiomycetes through a hollow fiber replica, and the cross-sectional area occupied by the hollow fiber membrane and the hollow part is 70% of the cross-sectional area of the container containing the hollow fiber membrane. 1. A method for cultivating mushrooms, characterized in that basidiomycetes are fully cultured on the outer wall surface of hollow fibers filled so as not to exceed 50%, and fruiting bodies are cultivated using the cultured mycelium. (2) The method according to claim 1, wherein the hollow fiber membrane has an average pore diameter of 300 μm to 2.0 μm.