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INVENTION
Patent of the Russian Federation RU2192119
A METHOD FOR PREPARING GRAIN MILITARY OF HIGHER MUSHROOMS
The name of the inventor: Kuznetsov O.Yu.
The name of the patent holder: Ivanovo State Medical Academy; Kuznetsov Oleg Yuvenalievich
Address for correspondence: 153462, Ivanovo, F. Engels ave., 8, IGMA, patent group
Date of commencement of the patent: 2000.05.06
(EN) The invention relates to biotechnology, in particular to methods for producing a cereal mycelium of higher (edible) fungi. The proposed method for growing the cereal mycelium of higher fungi is as follows: after filling the tanks with dry grain and water, the grain is soaked in water for 24 hours at room temperature to initiate spore germination of the lower fungi and transfer them to the vegetative state, and then drain the water. The wet grain is left in the containers, then the neck of the flexible unclosed on one side of the container is used to assemble the geometrically unchangeable system in such a way that the material of the open flexible container is between the components of the geometrically unchangeable system. At the same time, a sealing material separating the internal volume of the container from the surrounding medium and allowing the sterility of the grain in the container to be maintained on the inside of the flexible, non-closed container (as one of the components), and to ensure free gas exchange between the internal volume and the environment. The moistened grain is incubated for 24 hours under the same conditions until further autoclaving at a temperature of 121 ° C., 1 atm pressure for 2-2.5 hours. Next, the cereal tanks are cooled to room temperature and inoculated with the uterine mycelium and then, subject to aseptic rules Cut off the geometrically unchangeable system from the container, and the newly formed flexible unclosed on one side tightly packed and made in several places with punctures (sterile small diameter needle) to preserve the gas exchange of the inner capacity of the bag where the growing mycelium on the grains is located, with the surrounding environment. After sowing, the containers are incubated in a thermostat at a temperature of 24-26 ° C or at room temperature in special rooms on racks. After 7-10 days, the germinating mycelium in the containers is shaken to prevent the gluing of the grain and the activation of the growth of the mycelium. The method makes it possible to improve the reliability of grain sterilization, reduce the laboriousness of preparing the grain mycelium, and preserve its microbiological purity for a long time.
DESCRIPTION OF THE INVENTION
The invention relates to biotechnology, in particular to methods for preparing cereal mycelia of fungi for subsequent use in the cultivation of fruiting bodies of these fungi.
As a prototype, the method for preparing the cereal mycelium of higher fungi according to RF patent 2101914, 1998 (Kuznetsov O.Yu.), which is implemented in stages as follows.
1. Dry grain is packed in a container by 2/3 of their useful volume.
2. The containers with grain are filled with water in such a way that the water does not reach the edges of the containers by about 8-10 cm, because when the dry grain swells, its volume increases by about one and a half times.
3. Containers with grain are left for 24 hours to soak the grain and initiate sprouting spores of the lower fungi, i. E. Transfer from a stationary state to a vegetative phase of development. At the end of this time or later (a delay of 6-8 hours is possible) the water is drained through a sieve from the containers, and the grain is left moist in the containers for another day. At this stage, if necessary, various composite additives, for example chalk and gypsum, can be added thereto. After adding these additives, each container is shaken and the additives are evenly distributed throughout the container, and the neck of the container is covered with aluminum foil.
Thus, the grain in the tanks until the time of autoclaving should be a total of 48 hours in two phases - the water phase (soaking the grain in containers) - 24 hours and the wet phase (water is removed from the tanks, and the grain is incubated there another day) - 24 h .
4. The containers are placed in an autoclave and sterilized at a temperature of 121 ° C, a pressure of 1 atm (101.325 kPa) and a sterilization time of 2-2.5 hours, depending on the volume of sterilized grain in the containers (determined experimentally).
5. The containers with the grain are cooled to room temperature and then inoculated with uterine mycelium on a dense agarized nutrient medium from "sugar" tubes or Petri dishes.
6. After sowing, the containers are incubated in a thermostat at a temperature of 24-26 ° C or at room temperature in special rooms on racks.
7. After 7-10 days, containers with sprouted mycelium are shaken to prevent gluing of the grain and activating the growth of the mycelium.
This stage can be combined with the re-packing of grain from containers into polyethylene bags. Packages are tightly packed and made in several places by punctures with a sterile needle to preserve the gas exchange of the inner capacity of the bag, where the growing mycelium on the grains is located, with the surrounding environment. The packets are then placed in a refrigerator with a temperature of 2-4 o C. As a result of the development of the mycelium on the grain, self-heating in the mass of the mycelium proceeds to 24-26 o C and therefore the conditions of the refrigerator do not inhibit the growing mycelium and it gradually covers all the grains in its Package. This is facilitated by the fact that condensation forms inside the package, creating the necessary humidity.
8. The final readiness of the mycelium is determined visually - the mycelium grows and connects the grains to each other in a single solid white briquette. Before sowing in the mass of the prepared substrate, the mycelium is stored in a refrigerator at the same temperature of 2-4 ° C.
Disadvantages of this method is the use of aluminum foil, as a sealing material, covering the neck of the container, does not guarantee the preservation of sterility of the grain for a long time because of a loose fit of the foil to the neck of the container. In addition, when the grain is shaken in the container, it is possible to damage the foil with dense particles of the substrate (grains), followed by a violation of sterility.
The proposed method for preparing the cereal mycelium of edible fungi is realized in stages and in the following way.
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1. Dry grain is packed into flexible unclosed containers (see drawing). 2. The given containers with grain are filled with water in such a way that the water covers the grain and is above its edge by approximately 8-10 cm, because when the dry grain swells the latter increases its volume by about one and a half times. 3. Containers with grain are left for 24 hours filled with water to soak the grain and initiate sprouting spores of the lower fungi, i.e. Transfer from a stationary state to a vegetative phase of development. At the end of this time or later (a delay of 6-8 hours is possible), the water is drained from the containers, and the grain is left moist in the containers for another day. At this stage, if necessary, various composite additives, for example chalk and gypsum, can be added thereto. After adding these additives, each container is shaken and the additives are evenly distributed throughout the volume of the container. |
4. Then the flexible unclosed on one side is passed through this container through a geometrically unchangeable system (such as the Soviet Encyclopedic Dictionary, 1986, p. 291) in such a way that the material of an open flexible capacitance is between the components of a geometrically immutable system. At the same time, a sealing component of a geometrically immovable system separating the internal volume of the container from the environment and allowing to preserve the sterility of the grain in the container is used on the inside of the flexible open tank (an exemplary list of materials for the manufacture of the container, see Table 1), and to ensure free gas exchange between Internal volume and environment.
5. Containers with a geometrically immovable system are placed in an autoclave and sterilized in a mode that is determined by experiment, depending on the volume of sterilized grain in the containers.
6. Capacities with GNS containing the inside grain are cooled to room temperature, and then inoculated with uterine mycelium on a dense agarized nutrient medium from "sugar" tubes or Petri dishes through the STS, in accordance with aseptic rules.
7. After inoculation of the tanks with the STS, the STS is cut off from the tank with the observance of asepsis rules, and the newly formed flexible open on one side is tightly packed and punctured in a few places (sterile small diameter needle) to preserve the gas exchange of the inner capacity of the package where the growing mycelium On grains, with the environment.
8. After sowing, the containers are incubated in a thermostat at a temperature of 24-26 ° C or at room temperature in special rooms on racks.
9. After 7-10 days, the containers with sprouted mycelium are shaken to prevent gluing of the grain and activating the growth of the mycelium.
10. The packets are then placed in a refrigerator with a temperature of 2-4 o C. As a result of the development of mycelium on the grain, self-heating occurs in the mass of the mycelium to 24-26 o C and, therefore, the conditions of the refrigerator do not inhibit the growing mycelium and it gradually covers all the grains In your package. This is facilitated by the fact that condensation forms inside the package, creating the necessary humidity. The final readiness of the mycelium is determined visually - the mycelium grows and connects the grains to each other into a single whole white briquette. Before sowing in the mass of the prepared substrate, the mycelium is stored in a refrigerator at the same temperature of 2-4 ° C.
EXEMPLARY EMBODIMENT OF THE INVENTION
In order to successfully achieve the ultimate goal of preparing the cereal mycelium of higher fungi, it is necessary to perform step by step all the above operations, during which it is expected to use a flexible non-closed container on one side made of a heat-resistant material, for example a polypropylene film (melting point 170 o C) capable of withstanding different regimes Autoclaving to achieve the sterility of the grain inside it. A more complete list of materials for the manufacture of heat-resistant flexible non-closed capacity (see table).
Thus, the technical result of the proposed method is to increase the reliability of grain sterilization by placing it in a flexible open tank with a geometrically unchangeable system, which allows sterilization of the prepared grain directly in the container without a recharging step, which helps to prevent contamination of the grain in the container by foreign competing microorganisms At the overgrowth of grain in the container and thereby allows a long-term preservation of the microbiological frequency of the resulting cereal mycelium of higher edible fungi.
CLAIM
A method for preparing a cereal mycelium of higher fungi, comprising preparing the substrate by mixing the grain with water in a container followed by aging, removing excess water, adding additives, incubating and sterilizing, inoculating with the uterine mycelium, characterized in that the grain is placed in a flexible, unclosed on one side container, Which passes this side between components of a geometrically immovable system, where a sealing material is used on the inside of the flexible container at the place of formation of the geometrically unchangeable system, providing free gas exchange between the environment and the internal volume of the container while maintaining the sterility of the grain contained in the container.
print version
Date of publication 11.03.2007gg
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