Lactarius deliciosus hypha submerged fermentation culture medium and application thereof

Abstract

The invention relates to the technical field of lactarius deliciosus hypha submerged fermentation culture media, and discloses a lactarius deliciosus hypha submerged fermentation culture medium and application thereof.The lactarius deliciosus hypha submerged fermentation culture medium is prepared from, by weight, 10-15 parts of a carbon source, 5-10 parts of a nitrogen source, 20-25 parts of minerals, 15-20 parts of vitamins and 10-15 parts of a regulator, the nitrogen source is specifically yeast powder, peptone powder or amino acid, the mineral substance is specifically calcium, phosphorus, magnesium, potassium, iron, zinc, copper or manganese, the vitamin is specifically nicotinamide, pantothenic acid, riboflavin or vitamin B12, and the conditioning agent is specifically a pH buffer agent or a surfactant. The lactarius deliciosus hypha submerged fermentation culture medium and the application thereof have the advantages that the hypha growth speed and yield are increased, fermentation conditions are optimized, and the culture medium is suitable for large-scale production, and an effective tool and method are provided for production and application of lactarius deliciosus.

Description

Technical Field

[0001] The invention relates to the technical field of a submerged fermentation medium for lactarius mycelium, and in particular to a submerged fermentation medium for lactarius mycelium and application thereof. Background Art

[0002] Lactarius pine is a common edible fungus, and the mycelium deep fermentation medium can be used to cultivate and propagate the mycelium of Lactarius pine.

[0003] The existing culture medium for deep fermentation of Lactarius pine mycelium on the market may have the following defects and deficiencies

[0004] The costs are high, the nutritional composition is not precise, there is a lack of personalized regulation capabilities, there is a lack of stability and consistency, and information sharing is limited.

[0005] Therefore, we proposed a culture medium for deep fermentation of Lactarius pine mycelium and its application. Summary of the invention

[0006] 1. Technical issues to be resolved

[0007] In view of the shortcomings of the prior art, the present invention provides a deep fermentation culture medium for Pleurotus ostreatus mycelium and its application, which has the advantages of increasing the growth rate and yield of mycelium, optimizing fermentation conditions, and being suitable for large-scale production. It provides effective tools and methods for the production and application of Pleurotus ostreatus, and solves the problems of high cost and inaccurate nutritional components.

[0008] (II) Technical solution

[0009] To achieve the above object, the present invention provides the following technical solution: a submerged fermentation medium for Lactarius pine mycelium comprises the following raw materials in weight proportions: 10-15 parts of a carbon source, 5-10 parts of a nitrogen source, 20-25 parts of minerals, 15-20 parts of vitamins and 10-15 parts of a regulator.

[0010] The carbon source is specifically glucose, corn flour or starch, the nitrogen source is specifically yeast powder, peptone powder, or amino acid, the mineral is specifically calcium, phosphorus, magnesium, potassium, iron, zinc, copper or manganese, the vitamin is specifically nicotinamide, pantothenic acid, riboflavin or vitamin B12, and the regulator is specifically a pH buffer or a surfactant.

[0011] The application of the deep fermentation medium of Pleurotus ostreatus mycelium comprises the following steps:

[0012] 1) Preparation of culture medium: preparing a culture medium for deep fermentation of Lactarius pine mycelium according to a specific ratio. The composition of the culture medium can be adjusted as needed, including a carbon source, a nitrogen source, minerals, vitamins and regulators;

[0013] 2) Preparation of culture equipment: Prepare sterile culture equipment and sterilize it with high temperature and high pressure before use to ensure sterility;

[0014] 3) Inoculation: Inoculate the strain of Lactarius pine into a sterile culture medium, using sterile seed pieces or mycelium fragments for inoculation. Keep a sterile operating environment to avoid contamination by other bacteria;

[0015] 4) Culture condition control: Place the inoculated culture medium in a suitable environment, including suitable temperature, humidity and light conditions, with a temperature of 25-28 degrees Celsius and a high relative humidity;

[0016] 5) Cultivation observation and management: During the cultivation process, observe the growth of mycelium, including growth rate and mycelium morphology, and take corresponding management measures as needed, such as adjusting the culture conditions and preventing bacterial or fungal contamination;

[0017] 6) Mycelium harvesting: After the mycelium has grown to a certain extent, it can be harvested. The mycelium can be used to prepare the cultivation substrate for edible fungi, or for other further experiments or applications.

[0018] Preferably, in the 1) preparation of the culture medium, the components such as the carbon source, nitrogen source, minerals and vitamins to be used are determined according to the experimental requirements and the growth characteristics of Lactarius pine, and the carbon source, nitrogen source, minerals and vitamins are mixed together in a specific proportion according to the determined formula, and the mixed components are added to an appropriate amount of distilled water, and fully stirred and dissolved until the components are evenly distributed, and the pH value and concentration of the culture medium solution are adjusted according to the growth requirements of Lactarius pine, and the adjusted culture medium solution is poured into an appropriate amount of culture vessel, and sterilized using a high temperature and high pressure sterilizer.

[0019] Preferably, in the preparation of the culture vessels 2), suitable culture vessels are selected, such as culture bottles, test tubes, plates, and homogenization bottles, to ensure that different culture vessels can provide a suitable growth environment. Before high-temperature and high-pressure sterilization, the culture vessels need to be cleaned and disinfected first, and deionized water or chlorine-containing disinfectant is used to thoroughly clean and disinfect, ensuring that the surface of the vessel does not contain any bacteria, and the sterilization conditions including temperature, pressure and time parameters are determined. The sterilization temperature is 121° C., the sterilization time is 15 to 20 minutes, and the sterilization pressure is above 0.1 MPa. The culture vessel containing the culture medium is placed in a high-temperature and high-pressure sterilizer for sterilization. Before sterilization, the safety valve and exhaust valve of the sterilizer should be checked to be normal, and it should be ensured that there is no residual oxygen in the container to avoid substandard pressure or explosion.

[0020] Preferably, in the 3) strain inoculation, select good quality, pure and impurity-free strains, accurately prepare the Pleurotus ostreatus colony culture medium, add appropriate nutrients in strict accordance with the strain requirements, disinfect all instruments, reagents and culture media that come into contact with the strains, and ensure their sterility if sterile seed pieces or mycelium fragments are used. Move the seed pieces or mycelium fragments to the surface of the culture medium with a sterile pipette or spoon for inoculation. According to the growth characteristics of the strains and the requirements of the culture environment, control the inoculation density to avoid being too dense or too sparse. Perform the inoculation operation on a sterile operating table, and use sterile techniques, equipment and instruments to avoid secondary contamination.

[0021] Preferably, in the 4) culture condition control, the temperature range is 25-28 degrees Celsius, a humidity regulator is placed in the incubator to provide suitable humidity, the relative humidity is maintained at about 70-80%, an appropriate amount of indirect lighting or a low light environment is provided, direct sunlight is avoided from shining on the culture medium, the ventilation holes of the incubator are opened or a circulating fan is used to promote air circulation, the pH value of the culture medium is maintained between 5.5 and 7, and the pH value is regularly tested and adjusted to meet the needs of the strain.

[0022] Preferably, in the 5) cultivation observation and management, the growth of Pleurotus ostreatus mycelium is observed regularly, the growth rate and the range of mycelium extension are recorded, a ruler or a microscope can be used for measurement and observation, the morphological characteristics of mycelium, including color, texture, density and branching, are observed, and attention is paid to observe whether there are abnormal phenomena, such as mycelium discoloration, obvious wilting or spots. According to the observation results, the culture conditions are adjusted in time to promote the normal growth of mycelium, the aseptic operation principles are strictly followed, the sterile state of the culture vessel, culture medium and working environment is ensured, the culture vessel, workbench and instruments are kept clean, and disinfected thoroughly with disinfectant, the observation results, operation steps and experimental parameters are recorded in time, and effective data management is performed.

[0023] Preferably, in the 6) mycelium harvesting, the mycelium growth is observed, and the appropriate time is selected for harvesting after the mycelium reaches a certain extent. When harvesting the mycelium, sterile operation must be adopted to avoid bacterial or fungal contamination. The mycelium is carefully cut or scraped from the culture medium using sterilized scissors or scrapers and other tools to avoid damaging the mycelium structure. The operation is performed as gently as possible. The harvest amount of mycelium is reasonably controlled according to the experimental requirements and the amount of planting substrate. After the mycelium is harvested, it needs to be stored in an appropriate physical state, frozen, dried or directly transferred to the planting substrate. The information of mycelium harvesting, including harvesting time, harvesting amount, storage method, etc., is recorded in time. At the same time, important information such as strain name and harvesting date are marked on the harvesting container or bag.

[0024] (III) Beneficial effects

[0025] Compared with the prior art, the present invention provides a culture medium for deep fermentation of lactarius mycelium and its application, which has the following beneficial effects:

[0026] The invention discloses a deep fermentation medium for lactarius mycelia and an application thereof. The deep fermentation medium for lactarius mycelia contains rich nutrients such as carbon source, nitrogen source and trace elements, and can meet the needs of lactarius mycelia growth and metabolism. The deep fermentation medium provides suitable environmental conditions and nutrients, and can promote the rapid growth and expansion of lactarius mycelia to achieve a high yield. The design of the medium takes into account the characteristics of lactarius strains and the needs in the fermentation process. The fermentation conditions are optimized by adjusting parameters such as temperature, pH value and oxygen supply, so that the mycelia growth and metabolism are in an optimal state. The lactarius mycelia can be rapidly expanded and form a closely connected mycelia mesh structure by using the deep fermentation medium, so that the mycelia can be harvested quickly and efficiently. The deep fermentation medium is suitable for large-scale production, and can realize large-scale culture and application of lactarius mycelia by expanding the scale of a culture container or performing continuous fermentation. The deep fermentation medium has the advantages of increasing mycelia growth speed and yield, optimizing fermentation conditions, and being suitable for large-scale production, and provides an effective tool and method for the production and application of lactarius. DETAILED DESCRIPTION

[0027] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0028] Embodiment: The culture medium for deep fermentation of lactarius mycelium comprises the following raw materials in proportion by weight: 10-15 parts of carbon source, 5-10 parts of nitrogen source, 20-25 parts of minerals, 15-20 parts of vitamins and 10-15 parts of regulator.

[0029] The carbon source is specifically glucose, corn flour or starch, the nitrogen source is specifically yeast powder, peptone powder, or amino acid, the mineral is specifically calcium, phosphorus, magnesium, potassium, iron, zinc, copper or manganese, the vitamin is specifically nicotinamide, pantothenic acid, riboflavin or vitamin B12, and the regulator is specifically a pH buffer or a surfactant.

[0030] The application of the deep fermentation medium of Pleurotus ostreatus mycelium comprises the following steps:

[0031] 1) Preparation of culture medium: preparing a culture medium for deep fermentation of Lactarius pine mycelium according to a specific ratio. The composition of the culture medium can be adjusted as needed, including a carbon source, a nitrogen source, minerals, vitamins and regulators;

[0032] 2) Preparation of culture equipment: Prepare sterile culture equipment and sterilize it with high temperature and high pressure before use to ensure sterility;

[0033] 3) Inoculation: Inoculate the strain of Lactarius pine into a sterile culture medium, using sterile seed pieces or mycelium fragments for inoculation. Keep a sterile operating environment to avoid contamination by other bacteria;

[0034] 4) Culture condition control: Place the inoculated culture medium in a suitable environment, including suitable temperature, humidity and light conditions, with a temperature of 25-28 degrees Celsius and a high relative humidity;

[0035] 5) Cultivation observation and management: During the cultivation process, observe the growth of mycelium, including growth rate and mycelium morphology, and take corresponding management measures as needed, such as adjusting the culture conditions and preventing bacterial or fungal contamination;

[0036] 6) Mycelium harvesting: After the mycelium has grown to a certain extent, it can be harvested. The mycelium can be used to prepare the cultivation substrate for edible fungi, or for other further experiments or applications.

[0037] In the 1) culture medium preparation, according to the experimental requirements and the growth characteristics of Lactarius pine, the components such as the carbon source, nitrogen source, minerals and vitamins to be used are determined, and according to the determined formula, the carbon source, nitrogen source, minerals and vitamins are mixed together in a specific ratio, and the mixed components are added to an appropriate amount of distilled water, and fully stirred and dissolved until the components are evenly distributed, and the pH value and concentration of the culture medium solution are adjusted according to the growth requirements of Lactarius pine, and the adjusted culture medium solution is poured into an appropriate amount of culture vessels, and sterilized using a high-temperature and high-pressure sterilizer.

[0038] In the preparation of the culture apparatus 2), suitable culture apparatus are selected, such as culture bottles, test tubes, plates, and homogenization bottles, to ensure that different culture apparatus can provide a suitable growth environment. Before high-temperature and high-pressure sterilization, the culture apparatus needs to be cleaned and disinfected first. Deionized water or chlorine-containing disinfectant is used to thoroughly clean and disinfect the apparatus to ensure that the apparatus surface does not contain any bacteria. The sterilization conditions including temperature, pressure and time parameters are determined. The sterilization temperature is 121° C., the sterilization time is 15 to 20 minutes, and the sterilization pressure is above 0.1 MPa. The culture apparatus containing the culture medium is placed in a high-temperature and high-pressure sterilizer for sterilization. Before sterilization, the safety valve and exhaust valve of the sterilizer should be checked to see if they are normal, and it should be ensured that there is no residual oxygen in the container to avoid substandard pressure or explosion.

[0039] 3) In the bacterial strain inoculation, good quality, pure and impurity-free bacterial strains are selected, the Pleurotus ostreatus colony culture medium is accurately prepared, appropriate nutrients are added in strict accordance with the bacterial strain requirements, all instruments, reagents and culture media in contact with the bacterial strain are disinfected, if sterile seed bacterial sheets or mycelium fragments are used, their sterility must be ensured, the seed bacterial sheets or mycelium fragments are moved to the surface of the culture medium with a sterile straw or spoon for inoculation, the inoculation density is controlled according to the growth characteristics of the bacterial strain and the requirements of the culture environment to avoid being too dense or too sparse, the inoculation operation is performed on a sterile operating table, and sterile techniques, equipment and instruments are used to avoid secondary contamination.

[0040] In the above-mentioned 4) culture condition control, the temperature range is 25-28 degrees Celsius, a humidity regulator is placed in the incubator to provide suitable humidity, the relative humidity is maintained at about 70-80%, an appropriate amount of indirect lighting or a weak light environment is provided, direct sunlight is prevented from shining on the culture medium, the ventilation holes of the incubator are opened or a circulating fan is used to promote air circulation, the pH value of the culture medium is maintained between 5.5 and 7, and the pH value is regularly tested and adjusted to meet the needs of the strain.

[0041] 5) During the cultivation observation and management, the growth of the mycelium of Pleurotus ostreatus is observed regularly, and the growth rate and the range of mycelium extension are recorded. A ruler or a microscope can be used for measurement and observation, and the morphological characteristics of the mycelium, including color, texture, density and branching, are observed. Note whether there are abnormal phenomena, such as mycelium discoloration, obvious wilting or spots. According to the observation results, the culture conditions are adjusted in time to promote the normal growth of mycelium, the aseptic operation principle is strictly followed, the aseptic state of the culture vessel, culture medium and working environment is ensured, the culture vessel, workbench and instruments are kept clean, and disinfected thoroughly with disinfectant, the observation results, operation steps and experimental parameters are recorded in time, and effective data management is performed.

[0042] 6) During the mycelium harvesting, the mycelium growth is observed, and the appropriate time is selected for harvesting after the mycelium reaches a certain extent. When harvesting the mycelium, sterile operation must be adopted to avoid bacterial or fungal contamination. The mycelium is carefully cut or scraped from the culture medium using sterilized scissors or scrapers to avoid damaging the mycelium structure. The operation is performed as gently as possible. The harvest amount of mycelium is reasonably controlled according to the experimental requirements and the amount of planting substrate. After the mycelium is harvested, it needs to be stored in an appropriate physical state, frozen, dried or directly transferred to the planting substrate. The information of mycelium harvesting, including harvesting time, harvesting amount, storage method, etc., is recorded in time. At the same time, important information such as strain name and harvesting date is marked on the harvesting container or bag.

[0043] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A culture medium for deep fermentation of lactarius pine hyphae, characterized in that: The invention comprises the following raw materials in proportion by weight: 10-15 parts of carbon source, 5-10 parts of nitrogen source, 20-25 parts of minerals, 15-20 parts of vitamins and 10-15 parts of regulator.

2. The submerged fermentation medium for Pleurotus ostreatus mycelium according to claim 1, characterized in that: The carbon source is specifically glucose, corn flour or starch, the nitrogen source is specifically yeast powder, peptone powder, or amino acid, the mineral is specifically calcium, phosphorus, magnesium, potassium, iron, zinc, copper or manganese, the vitamin is specifically nicotinamide, pantothenic acid, riboflavin or vitamin B12, and the regulator is specifically a pH buffer or a surfactant.

3. The application of the submerged fermentation medium of Pleurotus ostreatus mycelium is characterized in that: The following steps are involved: 1) Preparation of culture medium: preparing a culture medium for deep fermentation of Lactarius pine mycelium according to a specific ratio. The composition of the culture medium can be adjusted as needed, including a carbon source, a nitrogen source, minerals, vitamins and regulators; 2) Preparation of culture equipment: Prepare sterile culture equipment and sterilize it with high temperature and high pressure before use to ensure sterility; 3) Inoculation: Inoculate the strain of Lactarius pine into a sterile culture medium, using sterile seed pieces or mycelium fragments for inoculation, and pay attention to maintaining a sterile operating environment to avoid contamination by other bacteria; 4) Culture condition control: Place the inoculated culture medium in a suitable environment, including suitable temperature, humidity and light conditions, with a temperature of 25-28 degrees Celsius and a high relative humidity; 5) Cultivation observation and management: During the cultivation process, observe the growth of mycelium, including growth rate and mycelium morphology, and take corresponding management measures as needed, such as adjusting the culture conditions and preventing bacterial or fungal contamination; 6) Mycelium harvesting: After the mycelium has grown to a certain extent, it can be harvested. The mycelium can be used to prepare the cultivation substrate for edible fungi, or for other further experiments or applications.

4. The use of the submerged fermentation medium for Pleurotus ostreatus mycelium according to claim 1, characterized in that: In the 1) culture medium preparation, according to the experimental requirements and the growth characteristics of Lactarius pine, the components such as the carbon source, nitrogen source, minerals and vitamins to be used are determined, and according to the determined formula, the carbon source, nitrogen source, minerals and vitamins are mixed together in a specific ratio, and the mixed components are added to an appropriate amount of distilled water, and fully stirred and dissolved until the components are evenly distributed, and the pH value and concentration of the culture medium solution are adjusted according to the growth requirements of Lactarius pine, and the adjusted culture medium solution is poured into an appropriate amount of culture vessels, and sterilized using a high-temperature and high-pressure sterilizer.

5. The use of the submerged fermentation medium of Pleurotus ostreatus mycelium according to claim 1, characterized in that: In the preparation of the culture apparatus 2), suitable culture apparatus are selected, such as culture bottles, test tubes, plates, and homogenization bottles, to ensure that different culture apparatus can provide a suitable growth environment. Before high-temperature and high-pressure sterilization, the culture apparatus needs to be cleaned and disinfected first. Deionized water or chlorine-containing disinfectant is used to thoroughly clean and disinfect the apparatus to ensure that the apparatus surface does not contain any bacteria. The sterilization conditions including temperature, pressure and time parameters are determined. The sterilization temperature is 121° C., the sterilization time is 15 to 20 minutes, and the sterilization pressure is above 0.1 MPa. The culture apparatus containing the culture medium is placed in a high-temperature and high-pressure sterilizer for sterilization. Before sterilization, the safety valve and exhaust valve of the sterilizer should be checked to see if they are normal, and it should be ensured that there is no residual oxygen in the container to avoid substandard pressure or explosion.

6. The use of the submerged fermentation medium of Pleurotus ostreatus mycelium according to claim 1, characterized in that: 3) In the bacterial strain inoculation, good quality, pure and impurity-free bacterial strains are selected, the Pleurotus ostreatus colony culture medium is accurately prepared, appropriate nutrients are added in strict accordance with the bacterial strain requirements, all instruments, reagents and culture media in contact with the bacterial strain are disinfected, if sterile seed bacterial sheets or mycelium fragments are used, their sterility must be ensured, the seed bacterial sheets or mycelium fragments are moved to the surface of the culture medium with a sterile straw or spoon for inoculation, the inoculation density is controlled according to the growth characteristics of the bacterial strain and the requirements of the culture environment to avoid being too dense or too sparse, the inoculation operation is performed on a sterile operating table, and sterile techniques, equipment and instruments are used to avoid secondary contamination.

7. The use of the submerged fermentation medium for Pleurotus ostreatus mycelium according to claim 1, characterized in that: In the above-mentioned 4) culture condition control, the temperature range is 25-28 degrees Celsius, a humidity regulator is placed in the incubator to provide suitable humidity, the relative humidity is maintained at about 70-80%, an appropriate amount of indirect lighting or a weak light environment is provided, direct sunlight is prevented from shining on the culture medium, the ventilation holes of the incubator are opened or a circulating fan is used to promote air circulation, the pH value of the culture medium is maintained between 5.5 and 7, and the pH value is regularly tested and adjusted to meet the needs of the strain.

8. The use of the submerged fermentation medium for Pleurotus ostreatus mycelium according to claim 1, characterized in that: 5) During the cultivation observation and management, the growth of the mycelium of Pleurotus ostreatus is observed regularly, and the growth rate and the range of mycelium extension are recorded. A ruler or a microscope can be used for measurement and observation, and the morphological characteristics of the mycelium, including color, texture, density and branching, are observed. Note whether there are abnormal phenomena, such as mycelium discoloration, obvious wilting or spots. According to the observation results, the culture conditions are adjusted in time to promote the normal growth of mycelium, the aseptic operation principle is strictly followed, the aseptic state of the culture vessel, culture medium and working environment is ensured, the culture vessel, workbench and instruments are kept clean, and disinfected thoroughly with disinfectant, the observation results, operation steps and experimental parameters are recorded in time, and effective data management is performed.

9. The use of the submerged fermentation medium of Pleurotus ostreatus mycelium according to claim 1, characterized in that: 6) During the mycelium harvesting, the mycelium growth is observed, and the appropriate time is selected for harvesting after the mycelium reaches a certain extent. When harvesting the mycelium, sterile operation must be adopted to avoid bacterial or fungal contamination. The mycelium is carefully cut or scraped from the culture medium using sterilized scissors or scrapers to avoid damaging the mycelium structure. The operation is performed as gently as possible. The harvest amount of mycelium is reasonably controlled according to the experimental requirements and the amount of planting substrate. After the mycelium is harvested, it needs to be stored in an appropriate physical state, frozen, dried or directly transferred to the planting substrate. The information of mycelium harvesting, including harvesting time, harvesting amount, storage method, etc., is recorded in time. At the same time, important information such as strain name and harvesting date is marked on the harvesting container or bag.