阅读说明：本技术 水杨酸或脯氨酸在香菇高温栽培中的应用 (Application of salicylic acid or proline in high-temperature cultivation of lentinus edodes ) 是由 赵媛 高淑敏 谭玉辉 段晓明 徐鸿雁 于 2020-05-08 设计创作，主要内容包括：本发明涉及食用菌培育技术领域,具体来说是水杨酸或脯氨酸在香菇高温栽培中的应用,本发明的用于香菇高温栽培的耐高温培养基由马铃薯培养基、浓度为10-50mg/L的水杨酸或浓度为10-50mg/L的脯氨酸组成,调节pH至7.2-7.4,本发明还公开了耐高温培养基在增强香菇耐高温性能的测试方法,本发明通过水杨酸或者脯氨酸来替代现有技术的外源物质,不仅增强了香菇的耐高温性能,而且水杨酸或者脯氨酸均廉价易得,从而有效降低了香菇菌种菌落培育的成本。(The invention relates to the technical field of edible mushroom cultivation, in particular to application of salicylic acid or proline in high-temperature cultivation of mushrooms, wherein a high-temperature-resistant culture medium for high-temperature cultivation of mushrooms comprises a potato culture medium, salicylic acid with the concentration of 10-50 mg/L or proline with the concentration of 10-50 mg/L, and the pH is adjusted to 7.2-7.4.)

1. The application of salicylic acid or proline in the high-temperature cultivation of shiitake mushrooms.

2. The high-temperature-resistant culture medium for the high-temperature cultivation of the shiitake mushrooms is characterized by comprising a potato culture medium, salicylic acid with the concentration of 10-50 mg/L or proline with the concentration of 10-50 mg/L, and the pH is adjusted to 7.2-7.4.

3. The thermostable culture medium for the hyperthermia cultivation of shiitake mushroom according to claim 2, wherein said culture medium is composed of potato culture medium, salicylic acid at a concentration of 25-50 mg/L or proline at a concentration of 10-25 mg/L, and the pH is adjusted to 7.2-7.4.

4. The thermophilic culture medium for the thermophilic culture of shiitake mushrooms as claimed in claim 2, wherein said medium consists of potato medium, salicylic acid at a concentration of 25 mg/L or proline at a concentration of 10 mg/L, adjusted to a pH of 7.2-7.4.

5. The method for testing the high-temperature-resistant culture medium for enhancing the high-temperature resistance of the shiitake mushrooms as claimed in any one of claims 2 to 4, which is characterized by comprising the following steps:

(1) strain activation

Inoculating the mushroom strain colony to a potato culture medium plate for activation treatment to obtain an activated strain;

(2) optimum concentration test

Inoculating the activated strain obtained in the step (1) to potato culture medium plates added with salicylic acid or proline with different concentrations, culturing at 25 ℃, and measuring the growth speed of hyphae by adopting a cross method;

wherein the concentration of the salicylic acid or the proline is 10-50 mg/L respectively;

(3) high temperature stress test

Inoculating the activated strain obtained in the step (1) to a potato solid culture medium added with salicylic acid or proline with different concentrations, continuously culturing at high temperature of 30 ℃, observing the growth condition of hyphae, and measuring the growth speed of the hyphae by adopting a cross method;

wherein the concentration of the salicylic acid or the proline is 10-50 mg/L respectively;

(4) cross high temperature stress test

Inoculating the activated strain obtained in the step (1) to a potato culture medium added with salicylic acid or proline with different concentrations, culturing at 25 ℃ in the dark for 5-7 days, and then placing at 40 ℃ in the dark for 24h for high-temperature stress; after high temperature stress, the culture medium is placed back to 25 ℃ for light-tight culture, and the growth rate of hyphae is measured by adopting a cross method;

wherein the concentration of the salicylic acid or the proline is 10-50 mg/L respectively.

6. The method as claimed in claim 5, wherein the activating treatment of step (1) is: inoculating the mushroom bacterial colony to a potato culture medium plate, culturing at 25 ℃ in the dark for 5 days with the hypha facing upwards.

7. A method for cultivating shiitake mushroom at a high temperature, comprising using the high temperature-resistant medium according to any one of claims 2 to 4.

Technical Field

The invention relates to the technical field of edible fungus cultivation, in particular to application of salicylic acid or proline in high-temperature cultivation of lentinus edodes.

Background

Temperature is one of the important environmental factors influencing the growth and development of mushroom L entula edodes (Berk.) Pegler, heat stress often causes the apoptosis and the decay of mushroom hyphae, reduces the resistance of mushroom hyphae to infectious microbes, causes serious economic loss, in addition, high temperature easily causes mushroom body deformity, secondary mushroom failure and the like, becomes a main problem of local serious yield reduction and even abstinence, so the high temperature resistance becomes the first index for evaluating the stress resistance of edible fungi.

During the long-term evolution of organisms, the high-temperature tolerance of the organisms is enhanced by means of synthesis of osmoregulation substances or induction of expression of antioxidant enzyme genes and the like, and further, the damage of high-temperature stress to the organisms is relieved. External application of salicylic acid has been shown to increase the tolerance of plants to high temperatures, but this effect varies with the concentration of salicylic acid treated. The study of the Pixie moth and the like shows that the low-concentration oyster mushroom mycelium has obvious promotion effect on the growth of the tested 3 strains of oyster mushroom mycelium, and the high-concentration oyster mushroom mycelium has inhibition effect. Researches on Liu Feng Min and the like show that salicylic acid reduces damage of high-temperature stress on the structure and the function of a plant cell membrane, reduces chlorophyll degradation and protects normal physiological functions of protein molecules by improving SOD activity and regulating osmotic pressure balance, so that the heat resistance of the blueberry is improved; studies of Yanghui and the like show that the addition of salicylic acid with higher concentration in the liquid culture process of the lentinus edodes can induce the production of more extracellular enzymes, thereby relieving the toxicity of cadmium to the hyphae of the lentinus edodes. Proline (Pro) is a small-molecule osmotic substance, is the most water-soluble amino acid, and can be accumulated in eubacteria, protozoa, marine invertebrates and plants under various stress conditions, and the accumulation of Pro in plants under the conditions of salt, drought, high and low temperature, heavy metal, germ invasion, anaerobism, insufficient nutrition, atmospheric pollution and UV irradiation is reported.

Disclosure of Invention

Aiming at the technical defects, the invention aims to provide the application of salicylic acid or proline in the high-temperature cultivation of the shiitake mushroom, the salicylic acid or the proline is used for replacing exogenous substances in the prior art, so that the high-temperature resistance of the shiitake mushroom is enhanced, and the salicylic acid or the proline is cheap and easy to obtain, so that the cost for cultivating the mushroom strain colony is effectively reduced.

In order to solve the above technical problems, a first object of the present invention is to provide an application of salicylic acid or proline in high temperature cultivation of shiitake mushrooms.

The second purpose of the invention is to provide a high-temperature-resistant culture medium for high-temperature cultivation of mushrooms, which consists of a potato culture medium, salicylic acid with the concentration of 10-50 mg/L or proline with the concentration of 10-50 mg/L, and the pH is adjusted to 7.2-7.4.

Preferably, the culture medium consists of potato culture medium, salicylic acid at a concentration of 25-50 mg/L or proline at a concentration of 10-25 mg/L, and the pH is adjusted to 7.2-7.4.

Preferably, the culture medium consists of potato culture medium, salicylic acid at a concentration of 25 mg/L or proline at a concentration of 10 mg/L, and the pH is adjusted to 7.2-7.4.

Based on the high-temperature-resistant culture medium, the invention also provides a test method of the culture medium for enhancing the high-temperature resistance of the shiitake mushrooms, which comprises the following steps:

(1) strain activation

Inoculating the mushroom strain colony to a potato culture medium plate for activation treatment to obtain an activated strain;

(2) optimum concentration test

Inoculating the activated strain obtained in the step (1) to potato culture medium plates added with salicylic acid or proline with different concentrations, culturing at 25 ℃, and measuring the growth speed of hyphae by adopting a cross method;

wherein the concentration of the salicylic acid or the proline is 10-50 mg/L respectively;

(3) high temperature stress test

Inoculating the activated strain obtained in the step (1) to a potato solid culture medium added with salicylic acid or proline with different concentrations, continuously culturing at high temperature of 30 ℃, observing the growth condition of hyphae, and measuring the growth speed of the hyphae by adopting a cross method;

wherein the concentration of the salicylic acid or the proline is 10-50 mg/L respectively;

(4) cross high temperature stress test

Inoculating the activated strain obtained in the step (1) to a potato culture medium added with salicylic acid or proline with different concentrations, culturing at 25 ℃ in the dark for 5-7 days, and then placing at 40 ℃ in the dark for 24h for high-temperature stress; after high temperature stress, the culture medium is placed back to 25 ℃ for light-tight culture, and the growth rate of hyphae is measured by adopting a cross method;

wherein the concentration of the salicylic acid or the proline is 10-50 mg/L respectively.

Preferably, the activation treatment method in step (1) is: inoculating the mushroom bacterial colony to a potato culture medium plate, culturing at 25 ℃ in the dark for 5 days with the hypha facing upwards.

The invention also provides a high-temperature cultivation method of the mushroom, which comprises the step of using the high-temperature resistant culture medium.

Compared with the prior art, the invention has the beneficial effects that:

1. in the early research, the invention discovers that the high temperature resistance of the mushroom hyphae can be improved by adding salicylic acid externally, the Salicylic Acid (SA) is a simple phenolic compound generated in a plant body and is considered as a plant endogenous signal molecule and a new plant hormone, and the fungal cell has a metabolic pathway similar to that of a higher plant; proline as a foreign substance has important application value in relieving damage to organisms caused by abiotic stress such as temperature stress, and the like, and the stress resistance research of proline at present mainly aims at plants as main research objects, but the research on the stress resistance of edible fungi is less. According to the research, the mushroom is taken as a material, and the influence of the exogenous salicylic acid or proline on the heat resistance, the biomass and an oxidation-oxidation resistance system of the mushroom hyphae under heat stress is researched by comparing the hyphae biomass before and after treatment of the exogenous salicylic acid or the exogenous proline, the antioxidant enzyme activity in organisms and the like, so that technical support is provided for establishing a mushroom high-temperature-resistant cultivation technical system.

2. The study showed that the biomass of Lentinus edodes in the potato medium with 25 mg/L salicylic acid added was 1.5 times that of comparative example 1 (blank control), the SOD activity in mycelia increased 3.12 times and more than 1.76 times that of comparative example 1 (blank control) after salicylic acid addition, the CAT activity in mycelia increased 2.01 times and 1.82 times that of 25 deg.C treatment after salicylic acid addition, and the POD activity after treatment was higher than that of comparative example 1 (blank control) under the same conditions.

When the concentration of salicylic acid reaches 25 mg/L, the colony diameter reaches 7.22mm after 5d of culture after the culture at the optimum concentration test of 25 ℃, the growth rate of mycelium after the culture at the high temperature stress test of 30 ℃ is 10.1 times of the growth rate of the comparative example 1 (blank control group), the growth rate of mycelium after the cross high temperature stress test is 4.6 times of the growth rate of the comparative example 1 (blank control group), and the salicylic acid is 25 mg/L in the potato culture medium added with salicylic acid, which is the optimum culture concentration.

As a result of investigation, the biomass of shiitake mushrooms in the potato medium supplemented with 10 mg/L proline was 0.6 times that in comparative example 1 (blank control), the SOD activity in mycelia after the addition of proline was 1.94 times that before the heat treatment, and the CAT activity in mycelia after the addition of proline was 1.21 times that before the heat treatment, and the POD activity after the treatment was higher than that in comparative example 1 (blank control) under the same conditions.

When the proline concentration reaches 10 mg/L, the colony diameter reaches 5.29mm after 5d of culture after the culture at the optimum concentration test of 25 ℃, the mycelium growth speed is 3.3 times of the growth speed of the comparative example 1 (blank control group) after the culture at the high temperature stress test of 30 ℃, the mycelium growth speed is 1.2 times of the growth speed of the comparative example 1 (blank control group) after the cross high temperature stress test, and the proline concentration is 10 mg/L proline in the proline-added potato culture medium at the optimum culture concentration.

Drawings

FIG. 1 shows the growth difference of the mycelium of Lentinus edodes cultured in the medium of salicylic acid with different concentrations at 25 ℃ for 5 days in examples 1 to 3 and comparative example 1;

FIG. 2 shows the growth difference of the mycelium of Lentinus edodes cultured in the medium of salicylic acid with different concentrations at 30 ℃ for 16 days in examples 1 to 3 and comparative example 1;

FIG. 3 shows that examples 1 to 3 and comparative example 1 of the present invention were cultured at 25 ℃ for 5 to 7 days in the absence of light and then cultured at 40 ℃ for 24 hours in the absence of light; then placing the mushroom to 25 ℃ for dark culture for 6 days, and adding salicylic acid culture media with different concentrations to culture the growth difference of the mushroom mycelium;

wherein ck in figures 1-3 is potato culture medium, 10 is potato culture medium containing 10 mg/L salicylic acid, 25 is potato culture medium containing 25 mg/L salicylic acid, and 50 is potato culture medium containing 50 mg/L salicylic acid;

FIG. 4 shows the growth difference of mushroom mycelia cultured in the culture media with different proline concentrations at 25 ℃ for 5 days according to the present invention in example 4 to example 6 and comparative example 1;

FIG. 5 shows the difference in the growth of the mycelia of Lentinus edodes cultured in the medium containing proline at different concentrations at 30 ℃ for 16 days according to examples 4 to 6 and comparative example 1 of the present invention;

FIG. 6 shows that examples 4 to 6 and comparative example 1 of the present invention were cultured at 25 ℃ for 5 to 7 days in the absence of light and then cultured at 40 ℃ for 24 hours in the absence of light; then putting the mushroom mycelia back to 25 ℃ for dark culture for 6 days, and adding proline culture media with different concentrations to culture the growth difference of the mushroom mycelia;

wherein ck in figures 4-6 is potato culture medium, 10 is potato culture medium containing 10 mg/L proline, 25 is potato culture medium containing 25 mg/L proline, and 50 is potato culture medium containing 50 mg/L proline.

Detailed Description

The following description of the preferred embodiments and accompanying fig. 1-6 are used in conjunction with the accompanying drawings in the embodiments of the invention to illustrate the preferred embodiments.

The mushroom strains 336093 used in the invention are all purchased from Beijing Beinanna Chuanglian union Biotechnology research institute;

salicylic Acid (SA) is a simple phenolic compound produced in plants, is considered as a plant endogenous signal molecule and a novel plant hormone and can improve the high-temperature resistance of the lentinus edodes, proline (Pro) is a micromolecular osmotic substance and is an amino acid with the largest water solubility, proline as a foreign substance has important application value in relieving the damage to organisms caused by abiotic stress such as temperature stress, the concentrations of the salicylic acid and the proline are selected and researched by the testing methods of the examples 1 to 6, wherein the potato culture media used in the examples 1 to 6 and the comparative example 1 are respectively composed of the following raw materials of 200g of potatoes, 20g of glucose, 15 to 20g of agar and 1000m L of distilled water.