A white Grifola frondosa strain, spores, fruiting bodies and their applications

Abstract

This invention provides a white Grifola frondosa strain, spores, fruiting bodies, and their applications. The Grifola frondosa strain, named Finc-G-W1, belongs to the Grifola frondosa family and is deposited at the Guangdong Provincial Microbial Culture Collection Center, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCCNO.62552 and deposit date of June 20, 2022. The white Grifola frondosa strain provided by this invention solves the problem of poor stability and uniformity of existing white Grifola frondosa varieties in factory-scale bottle cultivation.

Description

Technical Field

[0001] This invention belongs to the field of edible fungi, and relates to a white Grifola frondosa strain, spores, fruiting bodies and their applications. Background Technology

[0002] Grifola frondosa (Latin name: Grifola frondosa) is a rare edible and medicinal fungus belonging to the Basidiomycota, Agaricomycetes, Apophyceales, Polyporaceae, and Grifola frondosa genus. In China, it primarily grows on chestnut trees, its layered growth resembling chrysanthemums, and it emits a pleasant fragrance, hence its other names such as chestnut mushroom, lotus mushroom, thousand-Buddha mushroom, and cloud mushroom. Grifola frondosa possesses a unique taste, flavor, and rich nutritional and medicinal value, making it one of the most common types of mushrooms on the market.

[0003] Due to the extreme scarcity of wild maitake mushroom resources, since its medicinal properties were recognized, artificial cultivation has become an important way to meet the demand. Large-scale cultivation of maitake mushrooms in my country began in Qingyuan County, Zhejiang Province, and Qianxi County, Hebei Province. Based on production methods, maitake mushrooms can be divided into soil-covered cultivation and soilless cultivation, the latter including bag cultivation and bottle cultivation. Based on fruiting body color, they can be divided into three types: brown, gray, and white. Brown varieties are currently the most widely used type, followed by gray varieties, while white varieties are the rarest. White maitake mushroom varieties, also known as "white maitake" or "snow maitake," are highly popular in the market due to their unique appearance and have great development potential.

[0004] Currently, the diversity of cultivars of Grifola frondosa in China is limited. Most domestic enterprises use brown or gray varieties suitable for bag cultivation, and there are no high-quality white varieties. In particular, there is a lack of white varieties that still have good stability and consistency under the existing factory-scale bottle cultivation production conditions, which seriously restricts the economic benefits and development potential of large-scale factory production and commercial promotion. Summary of the Invention

[0005] The purpose of this invention is to provide a white marigold strain, spores, fruiting bodies, and their applications, aiming to solve the problem of poor stability and consistency of white marigold varieties in factory-scale bottle cultivation.

[0006] To address the aforementioned technical problems, this invention provides a white Grifola frondosa strain, named Finc-G-W1, belonging to the Grifola frondosa family. It is deposited at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC NO.62552 and deposit date of June 20, 2022.

[0007] The present invention also provides a white Grifola frondosa spore, which is a spore obtained after culturing the above-mentioned Finc-G-W1 strain.

[0008] The present invention also provides a white Grifola frondosa (monokinetic) protoplast, which is a protoplast obtained after culturing the above-mentioned Finc-G-W1 strain.

[0009] The present invention also provides a white Grifola frondosa mycelium, which is the mycelium obtained by culturing the above-mentioned Finc-G-W1 strain.

[0010] The present invention also provides a white grizzly fruiting body, which is the fruiting body obtained by cultivating the above-mentioned Finc-G-W1 strain.

[0011] The present invention also provides the application of the above-mentioned white marigold fruiting bodies in food processing.

[0012] The present invention also provides an application of the white Grifola frondosa strain in breeding, using Finc-G-W1 as a parent for breeding.

[0013] The present invention also provides an application of the white Grifola frondosa strain in hybridization breeding, using the Finc-G-W1 strain as a parent for hybridization breeding.

[0014] The present invention also provides an application of the white Grifola frondosa strain in breeding, wherein the Finc-G-W1 strain is subjected to molecular editing breeding.

[0015] Compared with the prior art, the white Grifola frondosa strain Finc-G-W1 provided by the present invention has the following advantages:

[0016] 1. The fruiting bodies cultured from the strain have good stability and reproducibility, and the differences in fruiting body traits between batches are small and the consistency is strong, which ensures the scale benefits of industrial production.

[0017] 2. The fruiting bodies are pure white, full in shape, fragrant, and have a good appearance;

[0018] 3. Short growth cycle, with an average cycle of ≤40 days, which improves the factory's production efficiency and effectively saves energy and reduces consumption;

[0019] 4. The fruiting bodies have a long shelf life; under 4℃ cold storage conditions, the average shelf life is ≥40 days, which is beneficial for long-distance transportation and extends the shelf life. Attached Figure Description

[0020] Figure 1 This is a morphological diagram of the fruiting body of the natural mutant Grifola frondosa strain F0 provided in this embodiment of the invention;

[0021] Figure 2 This is a conventional fruiting body morphology diagram of a normal Grifola frondosa strain provided in the embodiments of the present invention;

[0022] Figure 3This is a diagram of the hyphal morphology of the Finc-G-W1 strain in an embodiment of the present invention;

[0023] Figure 4 and Figure 5 This is a morphological diagram of the fruiting body of the Finc-G-W1 strain in an embodiment of the present invention. Detailed Implementation

[0024] The following detailed description, in conjunction with the accompanying drawings and specific embodiments, provides a further detailed account of the white Grifola frondosa strain, spores, fruiting bodies, and applications of the present invention. The advantages and features of the present invention will become clearer from the following description. It should be noted that the accompanying drawings are all in a very simplified form and use non-precise proportions, used only to facilitate and clarify the illustration of the embodiments of the present invention. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods. Unless otherwise specified, the materials and reagents used in the following embodiments are commercially available.

[0025] This invention provides a white Grifola frondosa strain, named Finc-G-W1, belonging to Grifola frondosa (Latin name: Grifola frondosa), deposited at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC NO.62552 and deposit date of June 20, 2022.

[0026] The Finc-G-W1 white Grifola frondosa strain of the present invention was obtained by continuous isolation and purification of a naturally mutated Grifola frondosa strain F0 during actual industrial production.

[0027] The present invention also provides a white Grifola frondosa spore, which is a spore obtained after culturing the above-mentioned Finc-G-W1 strain.

[0028] The present invention also provides a white Grifola frondosa (monokinetic) protoplast, which is a protoplast obtained after culturing the above-mentioned Finc-G-W1 strain.

[0029] The present invention also provides a white Grifola frondosa mycelium, which is the mycelium obtained by culturing the above-mentioned Finc-G-W1 strain.

[0030] This invention also provides the application of the aforementioned white Grifola frondosa mycelium in the extraction of active substances and secondary metabolites, and in fermentation engineering.

[0031] The present invention also provides a white grizzly fruiting body, which is the fruiting body obtained by cultivating the above-mentioned Finc-G-W1 strain.

[0032] This invention also provides the application of the above-mentioned white maitake fruiting bodies in food processing, such as the preparation of maitake polysaccharides, D-Fraction and other active substances.

[0033] The present invention also provides an application of the white Grifola frondosa strain in breeding, using Finc-G-W1 as a parent for breeding.

[0034] The present invention also provides an application of the white Grifola frondosa strain in hybridization breeding, using the Finc-G-W1 strain as a parent for hybridization breeding.

[0035] The present invention also provides an application of the white Grifola frondosa strain in breeding, wherein the Finc-G-W1 strain is subjected to molecular editing breeding.

[0036] like Figure 3 As shown, the white Grifola frondosa mycelium obtained by the Finc-G-W1 strain of the present invention is dense and white, with underdeveloped aerial mycelium, smooth colony surface, moderate thickness, no pigment, and relatively neat edges.

[0037] like Figure 4 and Figure 5 As shown, the fruiting bodies cultivated using the Finc-G-W1 strain of the present invention are full, expansive, densely packed, and arranged in an imbricate pattern. The fruiting bodies are white to milky white in color and have a fragrant aroma. The cap is fleshy, fan-shaped to spoon-shaped, with no obvious notches on the periphery, and the vertical section is nearly horizontal. The cap is white to milky white, with ridge-like protrusions on the surface, of medium thickness, and of medium hardness. The pores are attached to the back of the cap, extending to the base, and are white to milky white in color and of medium density. The stipe is fleshy, white to milky white, short to medium in length, medium in diameter, and thin to medium at the base.

[0038] To further highlight the advantages of the white Grifola frondosa strain provided by this invention, the following experiments provide a more detailed comparative explanation of the strain and its applications.

[0039] Example 1: Isolation and purification of Finc-G-W1 strain

[0040] In this embodiment, the specific breeding method of the white Grifola frondosa strain Finc-G-W1 is as follows:

[0041] The Finc-G-W1 white Grifola frondosa strain of this invention was isolated and purified from a naturally mutated Grifola frondosa strain F0 during actual factory production. Its fruiting body morphology is as follows: Figure 1 As shown, Figure 2 This represents the typical fruiting body morphology of a normal bacterial strain.

[0042] 1) Source of the mutant strain: A naturally mutated Grifola frondosa strain F0 from the actual factory production process.

[0043] 2) Prepare the culture medium (PDA medium): Use 200g of potato, 20g of glucose, and 20g of agar per 1L of culture medium.

[0044] Primary culture medium: comprising 35-40% primary culture medium and 60-65% water by weight percentage, wherein the primary culture medium comprises 80-90% sawdust and 10-20% wheat bran by weight percentage.

[0045] Cultivation medium: by weight percentage, it comprises 35-40% cultivation medium and 60-65% water. The cultivation medium comprises by weight percentage 20-25% chestnut sawdust, 20-25% oak sawdust, 15-20% cottonseed hulls, 10% wheat bran, 5-10% brewer's grains, and 5-10% soybean residue.

[0046] 3) Isolation and purification: The robust fruiting body stipes of the initial mutant strain F0 were selected as materials, and 80 pure cultures of its F1 generation PDA were obtained by tissue isolation and continuous transfer.

[0047] 4) Test tube preservation: Transfer the above F1 generation PDA pure culture into test tubes, and after the mycelium has fully expanded, place them in a cold storage at 4℃ for use. The storage time shall not exceed 3 months.

[0048] 5) Fruiting test: The F1 generation test tube culture was taken out and activated, transferred to a triangular flask, transferred to the original culture, transferred to the cultivation culture, cultured, scratched, and cultivated to produce fruiting.

[0049] 6) Fruiting screening: Select 10 F1 generation fruiting bodies with the lightest color based on the fruiting body color and production performance.

[0050] 7) Continuous isolation, purification and screening: Select the 10 F1 generation fruiting bodies with the lightest color and repeat the operation of steps 2)-6). From a total of 500 F2 generation fruiting bodies, screen out the target strain that is suitable for industrial bottle cultivation, has good stability and consistency, and has a pure white color and full flower shape. Number and name it "Finc-G-W1", abbreviated as "FG-W1".

[0051] 8) Strain preservation: The new white Grifola frondosa strain Finc-G-W1 of this invention was deposited at the Guangdong Provincial Microbial Culture Collection Center on June 20, 2022, with the accession number GDMCC NO.62552. The strain was found to be viable and its taxonomic position is Grifola frondosa.

[0052] Example 2: Cultivation Experiment

[0053] Multiple consecutive batches of fruiting experiments were conducted on the Finc-G-W1 strain of this invention at different times and seasons. Simultaneously, a comparative cultivation experiment was conducted using the commonly used Grifola frondosa strain MH as a control group. The cultivation experiment design in this embodiment involved six small-scale cultivation trials (320 bottles / trial), numbered as experimental batches 1, 2, 3, 4, 5, and 6.

[0054] The cultivation conditions used in this embodiment are as follows:

[0055] For the culture medium, please refer to the culture medium for cultivated varieties, which will not be repeated here.

[0056] Cultivation bottle volume: 850mL.

[0057] The LM culture medium formula is for cultivars (dry matter content: chestnut sawdust 20-25%, oak sawdust 20-25%, cottonseed hulls 15-20%, wheat bran 10%, brewer's grains 5-10%, soybean residue 5-10%; moisture content: 60%-65%).

[0058] Mycelial culture conditions: temperature 20-25℃, humidity 70-75%, intermittent diffused light, light intensity 50-100 lx, CO2 concentration controlled below 1500 ppm. The mycelial culture cycle is the number of days from inoculation to the presence of the fungus.

[0059] Fruiting body cultivation conditions: The cultivation room temperature should be set at 15-18℃, humidity at 85-95%, CO2 concentration controlled below 1200ppm, and light intensity at 50-100lx. Harvest the fruiting bodies promptly when they are close to maturity. The fruiting body cultivation cycle is the number of days from scratching the fungus to harvesting.

[0060] The fruiting data of the two strains are shown in Table 1 below.

[0061] Table 1 Comparison of fruiting data between strain F-G-W1 and strain MH

[0062]

[0063] As shown in Table 1, the culture period of FG-W1 strain remained stable at 23-24 days, the cultivation period at 13-14 days, and the total cycle at 36-38 days in the six batches of fruiting trials, which is 2-3 days shorter than that of MH strain used in production. Furthermore, the average yield of FG-W1 in the six batches of fruiting trials was more stable than that of MH, with each batch yielding an average of over 75g under these conditions. The culture period, cultivation period, and average yield of FG-W1 remained stable throughout continuous production, indicating good stability and repeatability. Moreover, the differences in fruiting bodies between batches and within each batch were small, demonstrating strong consistency.

[0064] Example 3: Preservation Test

[0065] In this embodiment, a preservation test was conducted on three batches of FG-W1 and MH. After harvesting, the whole fruit was quickly packaged, with 8 packages per batch of each material, and stored in a cold storage (4°C). The fruiting bodies were taken out once a week after the third week and the changes in fruiting body quality were recorded. The experimental results are shown in Table 2.

[0066] Table 2 Preservation Test Data

[0067]

[0068] As can be seen from the results in Table 2, the fruiting bodies obtained by strain FG-W1 have a longer shelf life than those obtained by strain MH, with an average shelf life of more than 45 days.

[0069] In summary, the Grifola frondosa strain Finc-G-W1 provided by this invention exhibits excellent overall characteristics and has broad market prospects, possessing the following advantages:

[0070] 1. The fruiting bodies cultured from the strain have good stability and reproducibility, and the differences in fruiting body traits between batches are small and the consistency is strong, which ensures the scale benefits of industrial production.

[0071] 2. The fruiting bodies are pure white, full in shape, fragrant, and have a good appearance;

[0072] 3. Short growth cycle, with an average cycle of ≤40 days, which improves the factory's production efficiency and effectively saves energy and reduces consumption;

[0073] 4. The fruiting bodies have a long shelf life; under 4℃ cold storage conditions, the average shelf life is ≥40 days, which is beneficial for long-distance transportation and extends the shelf life.

[0074] The above description is merely a description of preferred embodiments of the present invention and is not intended to limit the scope of the present invention in any way. Any changes or modifications made by those skilled in the art based on the above disclosure shall fall within the protection scope of the claims.

Claims

1. A type of white granola flower ( Grifola Frondosa strain Finc-G-W1, characterized in that, Grifola Frondosa, named Finc-G-W1, preserved in Guangdong Microbial Culture Collection Center, address: 59# Building, 5th Floor, Guangzhou Xianlie Middle Road 100 Courtyard, preservation number: GDMCC NO.62552, preservation date: June 20, 2022.

2. White Grifola frondosa spores, characterized in that, Spores obtained after culturing the white Grifola Frondosa strain Finc-G-W1 of claim 1.

3. A white Grifola frondosa protoplast, characterized by, Protoplasts obtained after culturing the white Grifola Frondosa strain Finc-G-W1 of claim 1.

4. A white Grifola frondosa mycelium, characterized in that, Mycelium obtained after culturing the white Grifola Frondosa strain Finc-G-W1 of claim 1.

5. A white Grifola frondosa fruit body, characterized by, Fruiting bodies obtained after culturing the white Grifola Frondosa strain Finc-G-W1 of claim 1.

6. The white Grifola Frondosa fruiting body of claim 5 for use in food processing.

7. Use of the white Grifola strain according to claim 1 for breeding, characterized in that, Breeding with the white Grifola Frondosa strain Finc-G-W1 as parent.

8. Use of the white Grifola strain according to claim 1 in crossbreeding, characterized in that, Cross breeding with the white Grifola Frondosa strain Finc-G-W1 as parent.

9. Use of the white Grifola strain according to claim 1 for breeding, characterized in that, Molecular editing breeding on the white Grifola Frondosa strain Finc-G-W1.

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