Black fungus strain Baiyun 9 and application thereof

Abstract

The invention belongs to the field of microorganisms, and relates to a black fungus strain Baiyun 9 and application thereof. The invention provides a novel black fungus strain which is obtained through brand new separation and successfully preserved, and the novel black fungus strain is rich in sporocarp nutrition, rich in various bioactive substances and capable of being well applied to the aspects of foods and medicines; a fresh product of the novel black fungus strain is ear-shaped, brown, semitransparent colloid, elastic, smooth and free of veins, and the average size of the novel black fungus strain is 5.6 * 3.8 cm; the hacking property is good, ear roots are not obvious, and the average thickness is 1.65 mm; after being dried, the product is strongly shrunk into cutin which is hard and crisp, the back surface is off-white and has short villus, and the ventral surface is black brown and smooth; after the dried agaric is fully soaked and loosened, the morphological characteristics of the dried agaric are almost the same as those of fresh agaric, and the soaking and loosening rate is not lower than 15.0; during stir-frying, a sound like setting off a firecracker can be made; the edible fungus culture medium has the advantages that the edible fungus culture medium is tender and crisp in taste, fresh, fragrant, glutinous and tasty, is deeply loved by people, is suitable for cultivation of various agriculture and forestry byproducts such as weed tree sawdust, mulberry branch sawdust, corncob and cottonseed hull, is short in fungus growing period and high in impurity resistance, and has good popularization and application and market prospects.

Description

Technical Field

[0001] The present application belongs to the field of microorganisms and relates to a black fungus strain Baiyun No. 9 and its application. Background Art

[0002] Black fungus (Auricularia auricular) belongs to the genus Auricularia of the phylum Eumycota, subphylum Basidiomycotina, class Hymenomycetes, order Auriculariales, family Auriculariaceae, kingdom Mycota, in fungal taxonomy. It is also known as black fungus, fungus, cloud fungus, etc.

[0003] Black fungus (Auricularia heimuer) is a typical colloid fungus, containing protein, dietary fiber, polysaccharides, amino acids, melanin, flavonoids, polyphenols, and 48 macro- and trace elements such as iron, zinc, calcium, and manganese. It is a nutritious edible fungus that is both medicinal and edible. Modern medical research has found that black fungus has multiple functions such as antioxidant and antibacterial activity and prevention of cardiovascular diseases. Actual clinical trials have proven that black fungus has the effect of preventing and treating coronary heart disease and atherosclerosis. Depending on the variety, origin, cultivation medium, and management method, the various nutrients and functional active substances contained in black fungus are different.

[0004] Black fungus produced in Guangxi is commonly known as cloud fungus. The main production area of ​​wild cloud fungus is in Baise. It belongs to the Auriculariales, Auriculariaceae, and Auricularia genus. It is a large colloid fungus that can be used for both medicine and food. As of the beginning of the 21st century, the production of Baise cloud fungus was still in a wild or semi-wild state, and its output was far from meeting the market demand. Therefore, separating and domesticating wild Baise cloud fungus and realizing artificial cultivation is the inevitable direction for the development of Baise cloud fungus.

[0005] With the development of biology and biotechnology, researchers hope to find more excellent black fungus strains with promotion and application value. Summary of the invention

[0006] In view of the shortcomings of the prior art, the present application provides a fresh black fungus strain, which has a bioconversion rate of not less than 130% and excellent appearance, and is an excellent black fungus strain with promotion and application value. The specific contents are as follows:

[0007] A black fungus strain is deposited in the General Microbiology Center of China Microorganism Culture Collection Committee with a deposit number of CGMCC No. 41269. The black fungus strain (Baiyun No. 9) is an excellent strain obtained by sampling from Baile Town, Tianlin County, Baise City, Guangxi Zhuang Autonomous Region, through separation, purification and qualitative domestication.

[0008] Furthermore, the black fungus strain includes spores, mycelium and fruiting bodies of the black fungus strain.

[0009] The application of the black fungus strain in food production.

[0010] The application of the black fungus strain in medicine preparation.

[0011] Furthermore, the black fungus strain includes spores, mycelium and fruiting bodies of the black fungus strain.

[0012] Fresh products are ear-shaped, brown, translucent, colloid, elastic, smooth, and without veins. Fig.14 ), good flaking, unclear ear roots, average size 5.6×3.8cm, average thickness 1.65mm; after drying, it shrinks strongly into keratin, hard and brittle, the back is grayish white with short hairs, the ventral side is dark brown and smooth; after the dried ears are fully soaked, their morphological characteristics are almost the same as those of fresh ears, and the soaking rate is not less than 15.0; when stir-fried, it will make a sound like setting off firecrackers; the taste is tender, crisp, fresh, fragrant, glutinous and refreshing, and is deeply loved by people. It is suitable for cultivation with a variety of agricultural and forestry by-products such as miscellaneous wood chips, mulberry branch chips, corn cobs (stalks), etc., with a short production cycle and strong resistance to weeds. The bioconversion rate of fresh ears is not less than 140%, and it has good promotion and application and market prospects.

[0013] The fresh fruiting body of the black fungus strain (Baiyun No. 9) described in the present application is ear-shaped, brown, translucent, colloid, elastic, smooth, and without veins, with an average size of 5.6×3.8 cm and an average thickness of 1.65 mm; after drying, it shrinks strongly into keratin, is hard and brittle, has a grayish white back with short hairs, and is dark brown and smooth on the ventral side; after the dried ear is fully soaked and loosened, its morphological characteristics are not significantly different from those of the fresh ear; when stir-fried, it will make a sound like setting off firecrackers; after cooking, it tastes smooth and not greasy, crisp and refreshing, and has a pleasant fragrance. It is an excellent black fungus strain with promotion and application value. It can be well applied to food and medicine.

[0014] The method for cultivating black fungus strains controls the temperature range to be 15-35° C. and the pH value range to be 5.5-6.5.

[0015] Furthermore, the temperature range is 25-28°C.

[0016] Furthermore, the pH value range is 6.0.

[0017] Furthermore, the water content of the solid culture medium used is 65%.

[0018] Furthermore, the culture medium used is as follows: 200 parts of potatoes (peeled), 20 parts of glucose, 3.0 parts of potassium dihydrogen phosphate, 1.5 parts of magnesium sulfate, 0.01 parts of vitamin B1, 1000 parts of water, and natural pH.

[0019] Beneficial effects of this application:

[0020] (1) This application provides a new strain of black fungus that was completely isolated and successfully preserved.

[0021] (2) The new black fungus strain provided in the present application has a fruiting body rich in nutrients and various biologically active substances. After cooking, it has a smooth and non-greasy taste, is crisp and refreshing, and has a pleasant fragrance, and can be well applied in food and medicine.

[0022] (3) The new strain of black fungus given in the present application is ear-shaped, brown, translucent, elastic, smooth, without veins, good at flake-opening, with inconspicuous ear roots, an average size of 5.6×3.8 cm, and an average thickness of 1.65 mm. After drying, it shrinks strongly into keratin, is hard and brittle, has a grayish white back with short hairs, and a dark brown and smooth ventral surface. After the dried ear is fully soaked and loosened, its morphological characteristics are not significantly different from those of the fresh ear, and it is crystal clear, bright and attractive, and has excellent market prospects.

[0023] (4) The new black fungus strain provided in the present application is an early-maturing variety. Compared with existing varieties, ear buds are formed after 10-15 days of mycelium culture ( Fig.15 ), no ripening period is required. Under normal circumstances, the fungus can be opened and grown after 35 days of cultivation, and the cultivation period is relatively shortened by 10-15 days. The ear-growing period is 40-60 days, which is 10-15 days shorter than that of other black fungus.

[0024] (5) The strain was cultured in pure form using mulberry twig particles (2-3 mm in diameter and 2-3 mm in length) and Cyclobalanopsis glauca particles (2-3 mm in diameter and 2-3 mm in length) and then stored in a refrigerator at 4-6°C for more than 13 years. There was no significant change in its performance indicators. However, there have been no reports of other varieties being preserved for a long time using this method. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a photo of the fresh fruiting body of Baiyun No. 9.

[0026] Figure 2 This is a photo of the dried fruiting body of Baiyun No. 9.

[0027] Figure 3 This is a photo of fresh fruiting bodies of Black 29.

[0028] Figure 4 Photos of the mycelium culture characteristics of Baiyun No. 9.

[0029] Figure 5 The electrophoresis patterns of various black fungus strains are as follows: 1-Baiyun No. 6, 2-Guiyun No. 3, 5-Baiyun No. 9, 6-Tianyun No. 1.

[0030] Figure 6 This is the developmental tree of each black fungus species, including: E1-Baiyun No. 6, E2-Guiyun No. 3, E5-Baiyun No. 9, E6-Tianyun No. 1.

[0031] Figure 7 This is the identification result of Baiyun No. 9 (E6), which was identified as a black fungus strain.

[0032] Figure 8 It is the antagonism line between Baiyun No. 9 and Tianyun No. 1.

[0033] Fig. 9 It is the antagonistic line between Baiyun No. 9 and Guiyun No. 3.

[0034] Fig.10 It is the antagonistic line between Baiyun No. 9 and Baiyun No. 6.

[0035] Fig.11 It is the antagonistic line between Baiyun No. 9 and Black 29.

[0036] Fig.12 It is the antagonistic line between Baiyun No. 9 and Hei Mu Er 916.

[0037] Fig.13 It is the confrontation line between Baiyun No. 9 and Heishan No. 15.

[0038] Fig.14 These are comparison photos of the appearance differences between the Baiyun No. 9 strain of the present application and the Hei No. 29 strain for comparison.

[0039] Fig.15 This is a photo showing ear buds formed after 10-15 days of culture of the mycelium of the Baiyun No. 9 strain of the present application. DETAILED DESCRIPTION

[0040] The present application is further described in detail below in conjunction with specific embodiments, which are intended to explain the present application rather than to limit it.

[0041] Example:

[0042] 1. Relevant information of the strain: including function, classification, properties, etc.

[0043] ①Classification status: Auricularia auricularia belongs to the genus Auricularia of the phylum Eumycota, subphylum Basidiomycotina, class Hymenomycetes, order Auriculariales, family Auriculariaceae, kingdom Mycota in fungal taxonomy. It is also known as black vegetable, fungus, cloud ear, etc.

[0044] ② Characteristics (including fruiting body morphology, mycelium culture characteristics, biological characteristics, cultivation characteristics, etc.):

[0045] a. Morphological characteristics of fruiting bodies Black fungus produced in Guangxi is commonly known as cloud fungus. The main production area of ​​wild cloud fungus is in Baise area. It belongs to the large colloid fungus of Auriculariales, Auriculariaceae, and Auricularia genus, which is used for both medicine and food. The Baiyun No. 9 of this application is an excellent strain with production and application value obtained by separation, purification, qualitative analysis, and domestication from the fruiting body samples collected from Baile Town, Tianlin, Baise, Guangxi at 106°13'\26°59'N\847m. The fresh fruiting body is ear-shaped, brown, translucent, colloid, elastic, smooth, without veins, good sliceability, and inconspicuous ear roots. The average size is 5.6×3.8cm and the average thickness is 1.65mm; after drying, it shrinks strongly into keratin, which is hard and brittle, with grayish white on the back and short villi, and dark brown and smooth on the ventral side; after the dried ear is fully soaked and loosened, its morphological characteristics are not significantly different from those of the fresh ear; when stir-fried, it will make a sound like setting off firecrackers; the taste is smooth and not greasy, crisp and refreshing, with a pleasant fragrance ( Figure 1 After drying, it shrinks strongly into keratin, which is hard and brittle. The back is off-white with short hairs, and the ventral side is dark brown and smooth. Figure 2 ); It is different from the main variety of black fungus in the main producing area of ​​Northeast China, Black 29, which is black, has obvious grid patterns (vein patterns, commonly known as "tendons") on the back (sterile side), and feels harder. Figure 3 ).

[0046] Baiyun No. 9 mycelium was cultured in a conventional comprehensive potato medium [200g potatoes (peeled), 20g glucose, 3.0g potassium dihydrogen phosphate, 1.5g magnesium sulfate, 10mg vitamin B1, 1000ml water, natural pH.] The culture characteristics are: round colonies, white, dense, strong and powerful mycelium, weak aerial mycelium, and basically no wall climbing phenomenon (lower Figure 4 ); identified by BGI as a black fungus variety (below Figure 5-7); It has a significant antagonistic reaction with the black fungus strains Hei 29, 916, Heishan 15, Baiyun 6, Guiyun 3, and Tianyun 1, which are currently applying for a patent. [Note: Individuals with significantly different genetic backgrounds will form a clear dividing line in the junction area, which is an antagonistic reaction. Antagonistic reaction is a specific manifestation of somatic cell incompatibility. It has been used in preliminary studies on fungal kinship because of its rapid, convenient and intuitive advantages. It was once the main means of fungal species identification. ] (below Figure 8-13 ), a new strain of black fungus.

[0047] The black fungus strain of the present invention is obtained by screening, domestication and cultivation of wild black fungus in Guangxi, and the screening and domestication methods are as follows:

[0048] (1) 30 samples of wild black fungus fruiting bodies collected from Baile Town, Tianlin, Baise, Guangxi (106°13'\26°59'N\847m) were used to obtain 21 pure strains through tissue separation and purification, and all of the 21 pure strains were identified as wild black fungus (cloud fungus) strains through direct ear extraction;

[0049] (2) A comparison test of ear growth was conducted on 21 wild black fungus (Auricularia auricula) strains. Based on the length of the mycelium maturity period and ear growth period, ear growth performance, ear piece color, the size of the ear root, the size and thickness of the ear piece, the ability to resist foreign matter, the foaming rate, and the bioconversion rate, the strain with a shorter mycelium maturity period and ear growth period, good ear growth performance, inconspicuous ear root, moderate and thick ear piece size, strong ability to resist foreign matter, and high foaming rate and bioconversion rate was selected as the mother strain Au-9;

[0050] (3) The mother strain Au-9 was cultivated in artificial bags and acclimated and screened in different seasons. Referring to the indicators in (2), 13 strains with better indicators, wide temperature adaptability and good high temperature resistance were selected and recorded as F1 generation;

[0051] (4) The F1 generation strains were cultivated in artificial bags and acclimatized and screened in different seasons. With reference to all the indicators in (2), 5 better strains were selected and recorded as the F2 generation;

[0052] (5) The F2 generation strains were cultivated in artificial bags and acclimatized and screened in different seasons. With reference to all the indicators in (2), three excellent strains were selected and recorded as the F3 generation;

[0053] (6) The F3 generation strains were cultivated in artificial bags and domesticated and screened in different seasons. Referring to all the indicators in (2), an excellent strain was selected with a mycelium maturity time of 35 to 40 days, a fast crop rotation, an ear-forming period of 40 to 60 days, good ear-forming performance, basically no ear roots, brown, translucent, elastic, an average ear size of 5.6×3.8 cm, an average thickness of 1.65 mm, a wide range of material suitability, strong resistance to impurities, a swelling rate of not less than 15%, and a fresh ear bioconversion rate of more than 140%. The strain was named "Baiyun No. 9" after the place of origin.

[0054] Among them, in step (1) to step (6), the tissue separation and purification method of the fruiting body is as follows: select a healthy and disease-free black fungus fruiting body, gently wipe the surface of the black fungus fruiting body with gauze soaked in 75% Vol alcohol solution, then put it into a sterilized culture dish and place it in a clean bench, turn on the ultraviolet lamp and the fan at the same time, and after 1 hour, when the moisture on the ear piece surface has dried and shrunk by 1\3 to 1\5, use tweezers and scissors burned by an alcohol lamp and cooled to cut a tissue block of about 2mm×2mm near the ear root, and insert it into

[0055] In a PAD slant medium without stagnant water, culture at a constant temperature of 25-28°C until the mycelium grows to 1\2-1\3 of the slant, pick the front mycelium, and transfer it to a fresh slant medium for culture. In the same way, continue to pick the front mycelium and transfer it to a fresh slant medium for culture. Repeat this method of selection and transplantation and combine microscopic examination until a pure strain is obtained. The formula of the test tube slant medium is: 1000ml of 20% peeled potato juice, 20g of glucose, 3g of potassium dihydrogen phosphate, 1.5g of magnesium sulfate, 15-17g of agar powder, and natural pH.

[0056] Among them, in step (1) to step (6), the black fungus cultivation and domestication method adopts bag-type cooked material cultivation, and the cultivation steps are as follows:

[0057] (1) Preparation of mushroom bags: Use 17cm×33cm×0.05mm low-density polyethylene edible fungus bags, put the culture material into the bag according to the conventional method, make mushroom bags, and then sterilize: When using normal pressure steam sterilization, the center material temperature should reach above 98℃ within 4 to 5 hours after the cold air is exhausted, and continue to maintain for 12 to 15 hours; when using high-pressure steam sterilization, the pressure reaches 0.15MPa after the cold air is exhausted, and the material temperature is maintained at 126℃ for 3 to 3.5 hours. After pre-cooling and forced cooling to a material temperature of 25 to 30℃, the sterilized mushroom bags are sent to the clean inoculation room and inoculated according to the requirements of the aseptic operation procedures.

[0058] The planting material consists of the following components in parts by weight: 0-78 parts of miscellaneous wood chips, 0-88 parts of mulberry branch chips, 0-90 parts of cottonseed hulls, 0-73 parts of corn cobs, 0-73 parts of bagasse, 7-15 parts of wheat bran, 0-3 parts of corn flour, 1-2 parts of soybean flour, 0-1 part of edible-grade glucose, 0.5-1 part of gypsum, and 0.5-1 part of lime.

[0059] (2) Mycelium culture: The inoculated fungus bags are promptly sent to the culture room and placed on shelves. The indoor environmental parameters are: temperature 25-28°C, humidity 55-65%, dim light, and fresh air. The culture time is 40-45 days.

[0060] (3) Opening the ear holes: The cultured mushroom bags (sticks) are transported to the place where the opening is to be made. The opening site should be clean and sanitary, disinfected with 0.1% potassium permanganate solution or carbendazim, and covered with a layer of clean plastic film. The opening device (machine) and the transport frame and other places that come into contact with the mushroom bag should also be disinfected. If an electric opening machine is used, care should be taken not to splash the solution onto the power supply and the motor. Then, 1 to 2 large barrels should be prepared, each containing about half a barrel of potassium permanganate solution for surface disinfection of the mushroom bag. When everything is ready, the mushroom bag opening operation can be officially started. Generally, three people are required to cooperate. One of them puts the mushroom bag to be opened into the disinfectant for surface disinfection, one person is responsible for opening, and the third person is responsible for receiving the bag and placing it in the transport frame. The mushroom bag is opened vertically, and the cutter wheel can be made of round nails, triangular nails, or straight nails. The hole spacing is 0.8 cm to 5 cm, the hole diameter is 0.3 cm to 0.6 cm, and the hole depth is 0.4 cm to 0.8 cm.

[0061] (4) Ear bud induction: Arrange the opened mushroom bags in a triangular shape on the ear-growing rack or on the surface of the ear-growing greenhouse, with a spacing of 15cm to 20cm between the mushroom bags and a shelf height of 45cm to 50cm. Management methods: ① Natural climate ear-growing greenhouse: Put down the film around the edges and close the door. Use the humidification equipment to automatically control the air humidity in the greenhouse to always be around 95%, the light intensity should not be less than 500lx, and the temperature and CO2 content in the greenhouse should be considered to determine whether to put down the film around the edges and close the door. Principle: If the greenhouse temperature exceeds 28℃, the film around the edges cannot be put down and the door can not be closed. Otherwise, the film around the edges can be put down and the door can be closed. ② Indoor ear-growing racks: Use relevant equipment to automatically control the indoor temperature to always be 25℃±2℃, the light intensity should not be less than 500lx, the air humidity should be around 95%, and the CO2 content should not be less than 500lx. 2 The content is within 500ppm. After 5-7 days of management, the primordium has basically begun to differentiate, and some may have formed ear buds. After 10-12 days, ear buds are formed, and more than 80% differentiate into ear pieces, and then enter the ear piece growth, which is the key water management stage.

[0062] (5) Management of ear growth: When most of the ear pieces grow to a size of 20mm, a sprinkler system should be used for water management. ① Natural climate ear growth greenhouse: When the temperature in the greenhouse is higher than 28℃, do not spray water. If the ear pieces are not dry, do not spray water. ② Indoor shelf ear growth: Use corresponding equipment to control the room to always maintain a temperature of 20-25℃ and clean air. When the dryness of the ear pieces reaches more than 80%, you can start spraying water.

[0063] The water temperature should be controlled between 20-25℃. The number of times and duration of water spraying depends on the state of the ear pieces. Each time, the ear pieces should be sprayed until they are fully expanded, like the state of dry wood ear fully soaked in water. Since dry wood ear pieces have a process of absorbing water, in order to save water, spray for 20-30 minutes, stop spraying for about 15 minutes, spray for another 20-30 minutes, and stop spraying for about 15 minutes. Repeat this 3-5 times until the ear pieces are fully expanded, and then wait for the next spray.

[0064] Follow the above principles to manage the ear pieces during their growth period until they are ready to be harvested. Under normal circumstances, this stage takes 10 to 15 days.

[0065] (6) Harvest at the right time

[0066] The fruiting bodies that have grown and matured should be harvested in time, and they should be harvested frequently and carefully to achieve the goal of high yield and high quality. The best harvesting period is when the ear pieces are 80% to 90% ripe. At this time, the ear pieces are of good quality and have weight. The standard for the maturity of wood ear is that the color turns light, the ear roots change from large to small, and the ear pieces stand upright. When the ear pieces are fully unfolded, the edges are wrinkled and thin, the ear roots shrink, and the back of the ear is light white, the spores are about to be ejected, and watering should be stopped before harvesting. The principle of picking ears is to harvest in time, pick the big ones and leave the small ones. If the ear pieces are not unfolded and have no commercial value, do not pick them, and keep them for the next harvest. Harvesting method: You can use a blade to cut them at the base of the ear without culture medium to keep the ear pieces clean, or you can hold the ear pieces by hand and twist them off at the base.

[0067] (7) Drying in time

[0068] After harvesting, the auricles should be dried in time. The methods include natural drying and hot air drying with drying equipment. The moisture content can be below 11%. Since Guangxi's special black fungus is rich in polysaccharides and pectin, it should be turned frequently during drying, especially at the beginning, to prevent the auricles from sticking together or sticking to the utensils. For the harvesting and drying method, a drying rack should be set up in advance. The rack should be 0.8 to 1.0 m above the ground, and a small pole should be placed on top. Screens and shade nets should be laid on top to facilitate ventilation. The natural light ventilation and cooling method not only improves the drying quality, but also solves the problem of fungus rotting on rainy days. Before drying, use scissors to cut off the culture medium at the root of the fungus. When drying, the leaves should be on top and the roots should be on the bottom. In addition, the clustered cloud ears must be torn into single pieces before drying. If it rains, cover it with rainproof material. The rainproof material should be covered above the drying rack, leaving a certain space for air circulation. Do not cover it directly on the ear pieces to prevent the ear pieces from being crushed and deformed, which will affect the quality of the fungus.

[0069] Finally, it should be pointed out that the above embodiments are only representative examples of the present application. Obviously, the technical solution of the present application is not limited to the above embodiments, and there may be many variations. All variations that can be directly derived or associated with the contents disclosed in the present application by a person of ordinary skill in the art should be considered as the protection scope of the present application.

Claims

1. A black fungus strain, characterized in that It is deposited in the General Microbiology Center of China Microorganism Culture Collection Administration with the deposit number CGMCC No.41269.

2. The black fungus strain according to claim 1, characterized in that The black fungus strain comprises spores, mycelium and fruiting bodies of the black fungus strain.

3. Use of the black fungus strain according to claim 1 in food production.

4. Use of the black fungus strain according to claim 1 in drug preparation.

5. The use according to claims 3 and 4, characterized in that: The black fungus strain comprises spores, mycelium and fruiting bodies of the black fungus strain.

6. The method for cultivating the black fungus strain according to claim 1, characterized in that: The controlled temperature range is 15-35°C and the pH value range is 5.5-6.

5.

7. The method for cultivating black fungus strain according to claim 6, characterized in that: The temperature range is 25-28°C.

8. The method for cultivating black fungus strain according to claim 6, characterized in that: The pH range is 6.

0.

9. The method for cultivating black fungus strain according to claim 6, characterized in that: The water content of the solid culture medium used was 65%.

10. The method for cultivating black fungus strain according to claim 6, characterized in that: The culture medium used was as follows: 200 parts of potatoes (peeled), 20 parts of glucose, 3.0 parts of potassium dihydrogen phosphate, 1.5 parts of magnesium sulfate, 1000 parts of water, pH natural.

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