Fungus with growth promoting function and preparation method and application of extracting solution of fungus

Abstract

The invention relates to the technical field of microorganisms, in particular to a preparation method and application of a fungus with a growth promoting function and an extract of the fungus, the classification name of the fungus is Fusarium Fraxinea (PK01), the preservation mechanism of the fungus is China General Microbiological Culture Collection Center (CGMCC), the preservation number is CGMCC No.42053, and the preservation date is July 9, 2025. Macrofungus sporocarp collected from Changbai Mountain wild is separated to obtain the fungus with a growth promoting function, the hypha extract under artificial culture conditions, and active ingredients of the hypha extract for promoting growth of main crops such as corn, improving drought tolerance and waterlogging tolerance, promoting growth of rhizosphere microorganisms, promoting flowering and improving yield.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to a fungus with growth-promoting function and a method for preparing and using its extract. Background Technology

[0002] Fungi are diverse and act as decomposers in ecosystems. They play an irreplaceable role in the development and evolution of the Earth's biosphere, the cycling of matter and energy, and the maintenance and restoration of the ecological environment. Many studies have shown that some fungi can form symbiotic relationships with plants, helping them absorb nutrients and enhance their resistance to adverse conditions. They can also be used to produce biopesticides to control crop diseases and pests. The isolation and extraction of these fungi has always been a major challenge. This application describes the *Fungiella ashensis* species *Bacillus perennialis*, which, through propagation, enables fungal extraction, and the extract promotes plant growth. Summary of the Invention

[0003] To address the aforementioned technical problems, this invention provides a method for preparing and using a fungus with growth-promoting function and its extract. The purpose is to promote crop root development, improve fertilizer utilization efficiency, enhance drought resistance, and increase yield through the application of related products.

[0004] The specific technical solution is as follows: Compared with the prior art, the present invention has the following beneficial technical effects: This invention isolates the fruiting bodies of large fungi collected from the wild in Changbai Mountain, obtaining fungi with growth-promoting functions, mycelial extracts under artificial culture conditions, and their active ingredients that promote the growth of major crops such as corn, improve drought and waterlogging resistance, promote rhizosphere microbial growth, promote flowering, and increase yield. The extracted active ingredients can be used in combination with a small amount of rhizosphere growth-promoting bacteria to treat corn and other crop seeds before sowing, and then sprayed on the crop leaves after flowering and pollination. This significantly promotes drought resistance and increases yield, greatly improving the economic benefits of crop cultivation. This invention also proposes a method for large-scale production of the fungal mycelium and extraction of its active ingredients using liquid aerobic fermentation technology, as well as field application techniques. Attached Figure Description

[0005] Figure 1 This is a plate diagram of fungal isolation according to the present invention; Figure 2 The effects of 10 ppb and 20 ppb fungal extracts on the germination of corn are shown in the figure. Figure 3 This is a liquid chromatogram of a fungal extract. Detailed Implementation

[0006] The following describes specific embodiments and accompanying drawings. Figures 1-3 The present invention will be described in detail, but the scope of protection of the present invention is not limited to the specific embodiments and drawings.

[0007] A fungus with growth-promoting function, the taxonomic name of which is: *Polyporus cremastrae* (PK01). Perenniporia fraxinea The depository is the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC No. 42053 and deposit date of July 9, 2025.

[0008] The amplified DNA sequence of this fungus is as follows: This sequence is the molecular basis for the classification of specialized types of *Fodium perfringens*. Example

[0009] A method for preparing a fungal extract with growth-promoting function, comprising the following steps: S1. Isolation of fungi: (1) Collect fungal fruiting bodies from under trees in the wild, place them in a collection box, and retrieve them. Immediately process and wash them, rinse the surface with 75% alcohol, wash with 10% pasteurized disinfectant for 10 minutes, and finally rinse twice with sterile water; (2) The above-disinfected fruiting bodies were cut into 0.5 cm long segments under aseptic conditions and inoculated onto Bengal red agar plates. After being placed in an incubator at 25°C for 4 days, mycelia were observed growing from the cut edges of the samples. The samples were then repeatedly separated and purified by plate culture. The sterilized fruiting bodies were aseptically cut into 0.5 cm segments and inoculated onto Bengal red agar plates. Hyphae were observed growing from the cut edges of the samples. The samples were repeatedly isolated and purified using the streak plating and dilution methods. The morphology of the colonies was observed, and the growth and characteristics of the colonies were recorded (including image recording). The strains were then numbered, and the samples proceeded to step S2, where the fungi were transferred to PDA slant plates and cultured for 4-9 days, then stored at 4°C. S2, Liquid fermentation culture of growth-promoting fungi: The fungi isolated in step S1 were inoculated onto PDA agar plates and cultured at 25°C for 7 days. Agar blocks were then inoculated into 250 mL Erlenmeyer flasks containing 100 mL of seed culture medium and cultured on a rotating shaker at 25°C and 150 r / min for 4 days as seed culture. The seed culture was then inoculated at 5% into 500 mL Erlenmeyer flasks containing 200 mL of fermentation medium and cultured under the same conditions for 7 days. Fermentation was then terminated and the samples were stored in a refrigerator at 4°C for later use. In step S2, the seed culture medium and the fermentation medium are the same, both being PDA medium; S3. Preparation of fungal mycelial alcohol extract: After washing the mycelium from step (2) above, dry it at 55°C, weigh it, and then pulverize it using a high-speed pulverizer. Extract it with the same volume of ethanol for 12 hours, repeat the extraction three times, mix it with a magnetic stirrer, ultrasonically vibrate it for 2 hours, vacuum filter it, and collect the filtrate for later use. Then, prepare the filtrate into a sample for testing and experimentation. Sample solution preparation: Dissolve the treated crude metabolite in an appropriate amount of methanol, filter it through an organic filter membrane with a pore size of 0.22 μm, discard the initial filtrate, and collect the subsequent filtrate as the sample solution. Example

[0010] A method for preparing a fungal extract with growth-promoting function, comprising the following steps: S1. Isolation of fungi: (1) Collect fungal fruiting bodies from under trees in the wild, place them in a collection box, and retrieve them. Immediately process and wash them, rinse the surface with 75% alcohol, wash with 10% pasteurized disinfectant for 15 minutes, and finally rinse three times with sterile water; (2) The above-disinfected fruiting bodies were cut into 0.5 cm long segments under aseptic conditions and inoculated onto Bengal red agar plates. After being placed in an incubator at 25°C for 9 days, mycelia were observed growing from the cut edges of the samples. The samples were then repeatedly separated and purified by plate culture. The sterilized fruiting bodies were aseptically cut into 0.5 cm segments and inoculated onto Bengal red agar plates. Hyphae were observed growing from the cut edges of the samples. The samples were repeatedly isolated and purified using the streak plating and dilution methods. Colony morphology was observed, and the growth and characteristics of the colonies were recorded (including image recording). The strains were then numbered, and the samples proceeded to step S2, where the fungi were transferred to PDA slant plates and cultured for 9 days before being stored at 4°C. S2, Liquid fermentation culture of growth-promoting fungi: The fungi isolated in step S1 were inoculated onto PDA agar plates and cultured at 25°C for 8 days. Agar blocks were then inoculated into 250 mL Erlenmeyer flasks containing 100 mL of seed culture medium and cultured on a rotating shaker at 25°C and 150 r / min for 4 days as seed culture. The seed culture was then inoculated at 5% into 500 mL Erlenmeyer flasks containing 200 mL of fermentation medium and cultured under the same conditions for 7 days. Fermentation was then terminated and the samples were stored in a refrigerator at 4°C for later use. In step S2, the seed culture medium and the fermentation medium are the same, both being PDA medium; S3. Preparation of fungal mycelial alcohol extract: After washing the mycelium from step (2) above, dry it at 55°C, weigh it, and then pulverize it using a high-speed pulverizer. Extract it with the same volume of ethanol for 12 hours, repeat the extraction three times, mix it with a magnetic stirrer, ultrasonically vibrate it for 2 hours, vacuum filter it, and collect the filtrate for later use. Then, prepare the filtrate into a sample for testing and experimentation. Sample solution preparation: Dissolve the treated crude metabolite in an appropriate amount of methanol, filter it through an organic filter membrane with a pore size of 0.22 μm, discard the initial filtrate, and collect the subsequent filtrate as the sample solution. Example

[0011] A method for preparing a fungal extract with growth-promoting function, comprising the following steps: S1. Isolation of fungi: (1) Collect fungal fruiting bodies from under trees in the wild, place them in a collection box, and retrieve them. Immediately process and wash them, rinse the surface with 75% alcohol, wash with 10% pasteurized disinfectant for 12 minutes, and finally rinse twice with sterile water; (2) The above-disinfected fruiting bodies were cut into 0.5 cm long segments under aseptic conditions and inoculated onto Bengal red agar plates. After being incubated at 25°C for 7 days, mycelia were observed growing from the cut edges of the samples. The samples were then repeatedly separated and purified by plate culture. The sterilized fruiting bodies were aseptically cut into 0.5 cm segments and inoculated onto Bengal red agar plates. Hyphae were observed growing from the cut edges of the samples. The samples were repeatedly isolated and purified using the streak plating and dilution methods. The morphology of the colonies was observed, and the growth and characteristics of the colonies were recorded (including image recording). The strains were then numbered, and the samples proceeded to step S2, where the fungi were transferred to PDA slant plates and cultured for 4-9 days, then stored at 4°C. S2, Liquid fermentation culture of growth-promoting fungi: The fungi isolated in step S1 were inoculated onto PDA agar plates and cultured at 25°C for 7 days. Agar blocks were then inoculated into 250 mL Erlenmeyer flasks containing 100 mL of seed culture medium and cultured on a rotating shaker at 25°C and 150 r / min for 4 days as seed culture. The seed culture was then inoculated at 5% into 500 mL Erlenmeyer flasks containing 200 mL of fermentation medium and cultured under the same conditions for 7 days. Fermentation was then terminated and the samples were stored in a refrigerator at 4°C for later use. In step S2, the seed culture medium and the fermentation medium are the same, both being PDA medium; S3. Preparation of fungal mycelial alcohol extract: After washing the mycelium from step S2 above, dry it at 55°C, weigh it, and then pulverize it using a high-speed pulverizer. Extract it with the same volume of ethanol for 12 hours, repeating the extraction three times. Mix the extract thoroughly with a magnetic stirrer, ultrasonically vibrate for 2 hours, and then vacuum filter it. Collect the filtrate for later use. Then, prepare the filtrate into a sample for testing. Sample solution preparation: Dissolve the treated crude metabolite in an appropriate amount of methanol, filter it through a 0.22 μm pore size organic filter membrane, discard the initial filtrate, and collect the subsequent filtrate as the sample solution.

[0012] Activity assay of the above fungal mycelial alcohol extract: The above-mentioned fungal mycelial alcohol extract was prepared into different concentrations. Corn seeds were soaked in the solution for 12 hours, and then placed into a germination machine according to different treatments. A 0.3% sodium chloride solution was prepared as the humidifying solution, and the seeds were cultured for 7 days according to the germination machine's preset program. Figure 3 The optimal concentration of the fungal extract for promoting corn germination is 20 ppb. The data above demonstrates that the fungal mycelium alcohol extract has a significant germination-promoting effect on corn seeds.

[0013] The application of fungal extracts in promoting maize growth and enhancing stress resistance is implemented as follows: Two plots of land not far apart were selected as comparative experimental fields, designated as the control group and the experimental group. The proximity of the two plots was designed to ensure a similar growing environment. Each plot was one acre in size, and the experimental sites were typical northern fields suitable for corn growth. Seeds of northern yellow corn were collected. The control group seeds were sown using conventional methods, while the experimental group seeds were treated with the fungal extract of this invention. The treatment steps were as follows: First, the corn seeds were soaked in the fungal extract at 10-20 ppb for 12 hours. The seeds from both plots were sown simultaneously. Second, the experimental group seeds were sprayed on the leaves approximately two weeks before flowering, or at the early stage of fruit set. Spraying was carried out on a sunny evening to avoid rain or sunlight affecting the crop's absorption of the spray. The standard for spraying was to ensure the leaves were evenly covered with the spray.

[0014] After reaching maturity, the two groups were observed separately. It was found that the maturity period of the control group was 135 days, while that of the experimental group was 125 days. This shows that the growth period of the corn in the experimental group was about ten days earlier than that of the control group. In terms of yield, the control group had a yield of nearly 1,400 jin per mu, while the experimental group had a yield of nearly 1,601 jin per mu. In other words, the experimental group using the fungal extract of this invention can significantly advance the maturity time of corn and increase the yield per mu.

[0015] This invention fully utilizes the relationship between fungi and plants, which can produce fungi that regulate plant growth substances, including beneficial and pathogenic fungi—a future technological trend. This invention isolates fungi from wild ginseng that promote the growth of crops such as corn. Analysis of the mycelial extract of this fungus revealed trace amounts of indoleacetic acid and gibberellin, as well as some unknown components. However, based on its very high activity, it is presumed not to be one of the five known classes of hormones. There have been no previous reports on similar fungi, especially considering that these active ingredients, in addition to promoting root growth, also induce drought and low-temperature tolerance in plants and promote the growth of rhizosphere microorganisms. The fungi of this invention can evolve symbiotically with plants to promote plant growth and development. During this symbiotic evolution, a mutually beneficial mechanism has been formed, which will gradually become clear through analysis of the active ingredients produced by the fungi. Artificially producing large quantities of fungi to obtain beneficial crop growth substances is an economical, effective, and environmentally friendly industry.

Claims

1. A fungus with growth-promoting function, characterized in that, The fungus is classified as: *Polyporus cremastrae* (PK01). Perenniporia fraxinea The depository is the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC No. 42053 and deposit date of July 9, 2025.

2. A fungus with growth-promoting function according to claim 1, characterized in that: The amplified DNA sequence of this fungus is as follows: , This sequence is the molecular basis for the classification of specialized types of *Fodium perfringens*.

3. The method for preparing the fungal extract with growth-promoting function as described in claim 1, characterized in that, The steps are as follows: S1. Isolation of fungi: (1) Collect fungal fruiting bodies under trees in the wild, put them in a collection box, and retrieve them; immediately process and wash them, rinse the surface with 75% alcohol, wash with 10% pasteurized disinfectant for 10-15 minutes, and finally rinse with sterile water 2-3 times. (2) The above-disinfected fruiting bodies were cut into 0.5 cm long segments under aseptic conditions and inoculated onto Bengal red agar plates. After being placed in an incubator at 25°C for 4 to 9 days, mycelia were observed growing from the cut edges of the samples. The samples were then repeatedly separated and purified by plate culture. S2, Liquid fermentation culture of growth-promoting fungi: The fungi isolated in step S1 were inoculated onto PDA agar plates and cultured at 25°C for 7-8 days. Agar blocks were then inoculated into 250 mL Erlenmeyer flasks containing 100 mL of seed culture medium and cultured on a rotating shaker at 25°C and 150 r / min for 4 days as seed culture. The seed culture was then inoculated at 5% into 500 mL Erlenmeyer flasks containing 200 mL of fermentation medium and cultured under the same conditions for 7 days. Fermentation was then terminated and the fungi were stored in a refrigerator at 4°C for later use. In step S2, the seed culture medium and the fermentation medium are the same, both being PDA medium; S3. Preparation of fungal mycelial extract: After washing the mycelium in step S2 above, dry it at 55°C, then pulverize it with a high-speed pulverizer. Weigh 10 grams of the dried mycelium, extract it with 100 ml of analytical grade ethanol for 12 hours, and repeat the extraction 3 times. Mix it with a magnetic stirrer, then vibrate it with ultrasound for 2 hours. Finally, filter it under vacuum with a filter membrane with a pore size of 0.22 μm and collect the fungal extract for later use.

4. The use of the growth-promoting fungal extract as described in claim 3, characterized in that: The extract was applied to field crops to promote crop growth, improve stress resistance, and increase yield. The specific usage process is as follows: The preferred method is to soak field crop seeds in a fungal extract at 10-20 ppb for 12 hours. Secondly, spray the leaves of field crops with bacterial solution two weeks before flowering, or spray the leaves of crops in the early stage of fruit setting. The application of fungal extract should be carried out on a sunny evening to avoid rain or sunlight affecting the crop's absorption of the sprayed solution. The standard for spraying is to ensure that the leaf surface is evenly covered with the sprayed solution.

Images