CN108432547B - Synthetic culture method for mycorrhiza - Google Patents
Synthetic culture method for mycorrhiza Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G18/00—Cultivation of mushrooms
- A01G18/10—Mycorrhiza; Mycorrhizal associations
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Abstract
The invention discloses a mycorrhiza synthetic culture method, which comprises the steps of culturing disinfected seeds or tissue culture seedlings in an isolated space in sterile nutrient solution, replacing the nutrient solution with target mycorrhizal fungi-containing bacterial liquid after lateral roots grow out of the seedlings, and promoting the root system and the target fungi to establish symbiotic relationship to form mycorrhiza, wherein the mycorrhiza is synthesized by means of early-stage sterile nutrient solution culture and later-stage replacement of the target bacterial liquid, so that the synthetic infection rate of the mycorrhiza is ensured to be more than 95%, and the large-scale production of the mycorrhiza seedlings is realized; the operation method is simple and convenient, has low cost and is suitable for large-scale production and application. The invention is suitable for mycorrhiza synthesis.
Description
Technical Field
The invention belongs to the field of agriculture, and relates to a mycorrhiza synthetic culture method.
Background
Mycorrhizal symbiosis, i.e. symbiosis formed by some fungi and plant roots in mycorrhizal soil, is widely distributed, and more than 80 percent of terrestrial plants can form symbiosis with mycorrhizal fungi. The fungi absorb organic substances such as carbon sources from plants as their own nutrients, and on the other hand, expand the root absorption surface, absorb nutrients (nutrient elements such as phosphorus and nitrogen in the soil) from the soil, supply water to the plants, and remarkably promote the growth of the plants.
The fungus symbiotic with the plant root system is called mycorrhizal fungi, the mycorrhizal fungi expand the nutrition absorption area of the plant root system, and the mycelial bridge enables the forest in the forest land to become a life and transport community of required moisture and nutrient substances. The mycorrhiza can not only promote the plant to absorb trace elements such as phosphorus, zinc, copper, calcium, magnesium, iron, manganese, boron and the like, but also generate auxin to promote the plant growth, enhance the disease resistance of the plant, improve the rhizosphere environment of the plant, improve the soil structure and promote the plant growth. In recent researches, there are some methods related to mycorrhiza synthesis, and all the methods have certain defects: such as complicated operation and no mass production; or the mycorrhiza synthesis rate cannot be effectively ensured, and the pollution cannot be effectively controlled in the implementation process.
Disclosure of Invention
The technical problem to be solved by the invention is to provide a mycorrhiza synthesis culture method, which is simple and convenient, easy to control the process, low in production cost and high in mycorrhiza synthesis rate.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows:
a mycorrhiza synthesis culture method is sequentially carried out according to the following steps:
(1) preparing a target plant growth nutrient solution and performing sterilization treatment, marked as A, and performing disinfection and sterilization treatment on a seedling culture container in the isolated space, marked as B;
the target plant growth nutrient solution consists of a fertilizer dissolved in water, the main components of the nutrient solution comprise nitrogen, phosphorus, potassium, calcium, magnesium, sulfur and a plurality of trace elements necessary for plant growth, and the nutrient solution is prepared according to the nutrition required by different plants;
(2) cultivating the disinfected seeds or tissue culture seedlings in the solution B containing the solution A;
(3) maintaining the circulation flow of the A, sterilizing the A flowing out of the B and then injecting the A into the B;
(4) preparing target bacterium liquid marked as C;
(5) gradually adjusting the pH value of A to be consistent with the pH value suitable for growth of C;
(6) after lateral roots grow out of the seedlings, replacing A with C, and culturing for 15 days to 3 months under the illumination condition to obtain the target mycorrhiza.
As a limitation of the present invention:
in the step (1), the temperature in the isolation space is 8-28 ℃, the isolation space keeps ventilation and light transmission, and the air in the isolation space is filtered by an air filter to prevent the pollution of mixed bacteria;
in the step (2), submerging the disinfected seeds in the A;
in the step (2), the tissue culture seedling is obtained by thoroughly cleaning the root with sterile water after the rooting culture is finished and before the seedling is hardened, and the root system of the tissue culture seedling is submerged in the A.
In the step (6), the cultivation is carried out under the illumination condition, the height of the liquid level is adjusted periodically, and part of the root system is exposed in the air, so that the respiration of the root is facilitated, and the oxygen deficiency of the root is avoided.
Due to the adoption of the technical scheme, compared with the prior art, the invention has the technical progress that:
the invention effectively controls the pollution possibly caused by the host plant in the processes of germination and growth (such as watering, temperature control and other actions in the production process) in the production process, saves the production cost through liquid culture, reduces the verbosity of the production link, and fully considers the growth cycle and the biological activity of the host plant and the target fungus hypha: after lateral roots grow out from the plant, the plant and the target bacteria are synthesized into mycorrhiza, and the mycorrhiza synthesis rate of the target bacteria is ensured to reach more than 95% by adjusting the pH value and periodically replacing bacterial liquid; the preparation method is simple, practical and efficient, has low production cost, is not influenced by seasons, and has remarkable economic benefit.
The invention is suitable for mycorrhiza synthesis.
The present invention will be described in further detail with reference to specific examples.
Detailed Description
The methods described in the following examples are those which are conventionally used unless otherwise specified, and the reagents used are those which are conventionally used unless otherwise specified.
Example 1A synthetic culture method for boletus edulis and Yunnan pine mycorrhiza
The embodiment is a method for synthesizing and culturing boletus edulis and mycorrhiza of Yunnan pine, which is carried out according to the following steps:
1) separating flesh from the boletus edulis sporocarp by adopting a tissue separation method, inoculating the flesh on a PDA (personal digital Assistant) plate culture medium, placing the culture medium in an incubator to culture and sprout hyphae at the temperature of 22-24 ℃, and transferring the hyphae to a new PDA plate culture medium again for purification culture to obtain pure hyphae;
2) after the hyphae grow over the culture dish, selecting the hyphae on an improved MMN liquid culture medium, and placing the hyphae in a shaking table for liquid fermentation culture for one month;
3) the greenhouse is improved, an air filtering system is added, ventilation and light transmission are kept, no mixed bacteria pollution exists in the air, and the temperature in the greenhouse is controlled to be 8-28 ℃;
4) preparing a Yunnan pine liquid nutrient solution and carrying out sterilization treatment, wherein each 100L of water contains 51g of urea, 31g of monopotassium phosphate, 10g of calcium sulfate, 5 g of magnesium sulfate, 0.01g of zinc sulfate, 0.03g of ferric sulfate, 0.01g of copper sulfate, 0.03g of manganese sulfate and 0.02g of boric acid powder;
5) sterilizing the surface of Yunnan pine seeds by using diluted potassium permanganate solution;
6) sterilizing facilities and equipment in the greenhouse by using ozone, placing the seeds sterilized in the step 5) into a seedling raising device, and injecting prepared Yunnan pine liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
7) keeping the nutrient solution in the seedling raising device to circularly flow, simultaneously sterilizing the nutrient solution flowing out of the seedling raising device by using an ultraviolet overflowing type sterilization system, and injecting the sterilized nutrient solution into the seedling raising device;
during the culture period, the height of the liquid level is controlled every day to ensure that the newly germinated Yunnan pine root system can contact the air every day;
8) measuring the pH value of the nutrient solution by using a pH meter, and adjusting the pH value to 6.0 by using diluted hydrochloric acid or sodium hydroxide;
9) after 3 lateral roots grow out from the Yunnan pine, closing the overflowing type sterilization system, and replacing the Yunnan pine liquid nutrient solution with boletus liquid diluted by 800 times;
during the culture period of the boletus liquid, the height of the liquid level is controlled every day, and the root system of the newly germinated Yunnan pine can contact the air every day; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
10) after culturing for 2 months under the illumination condition, the boletus edulis mycorrhiza is obtained.
The mycorrhiza synthesis rate of the embodiment reaches 98%, the preparation method is simple, practical and efficient, the production cost is low, the influence of seasons is avoided, and the economic benefit is remarkable.
Example 2A synthetic culture method for mycorrhiza of Thelephora ganbajun and Sequoia yunnanensis
The embodiment is a mycorrhiza synthesis culture method of Thelephora ganbajun and Sequoia yunnanensis, which is carried out according to the following steps:
1) separating spores of the sparassis crispa sporocarp by adopting a spore separation method, inoculating the spores on a PDA (personal digital Assistant) plate culture medium, placing the spores in an incubator to culture and sprout hyphae at the temperature of 22-24 ℃, and transferring the hyphae to a new PDA plate culture medium again for purification culture to obtain pure hyphae;
2) after the hyphae grow over the culture dish, picking the hyphae on an MMN liquid culture medium, and placing the MMN liquid culture medium in a shaking table for liquid fermentation culture for one month;
3) modifying a glass greenhouse, adding a fresh air system capable of isolating bacteria, keeping ventilation and light transmission, and controlling the temperature in the glass greenhouse to be 8-28 ℃, wherein the air is free from mixed bacteria pollution;
4) preparing a cryptomeria fortunei liquid nutrient solution and carrying out sterilization treatment, wherein each 100L of water contains 11g of sodium nitrate, 71 g of calcium superphosphate, 25 g of ammonium sulfate, 35 g of potassium sulfate, 42 g of magnesium sulfate, 2.4 g of ferrous sulfate, 0.35 g of boric acid powder, 0.25 g of manganese sulfate and 0.2g of trace elements;
5) sterilizing the surface of the Yunnan oil fir seeds by using 75 percent alcohol;
6) spreading lime and ground cloth on the ground in the glass greenhouse to prevent the propagation of fungi, performing spray sterilization on facilities and equipment by using 75% alcohol, placing the sterilized seeds obtained in the step 5) in a glass container, and injecting a yunnan fir liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
7) maintaining the circulation flow of the Yunnan fir liquid nutrient solution in the glass container, filtering and sterilizing the nutrient solution flowing out of the container by a filtering method, and injecting the sterilized nutrient solution into the glass container;
during the culture period, the liquid level of the nutrient solution is controlled to ensure that the newly germinated oil fir root system can contact the air every day;
8) measuring the pH value of the nutrient solution by using a pH meter, and adjusting the pH value to 6.0 by using diluted hydrochloric acid or sodium hydroxide;
9) after 2 lateral roots of the Yunnan oil fir grow out, closing a filtering and sterilizing system, and replacing the nutrient solution with thelephora ganbajun bacterial solution diluted to 1000 times;
after the rhizopus chinensis liquid is replaced by the rhizopus chinensis liquid, the height of the liquid level is controlled, and the root system of the newly germinated cryptomeria yunnanensis can be ensured to contact the air every day; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
10) after 2 months of culture, the dry mycorrhiza of the Yunnan oil fir is obtained.
The mycorrhiza synthesis rate of the embodiment reaches 97%, the preparation method is simple, practical and efficient, the production cost is low, the influence of seasons is avoided, and the economic benefit is remarkable.
Example 3A synthetic culture method of boletus edulis and chestnut mycorrhiza
The embodiment is a method for synthesizing and culturing boletus edulis and Chinese chestnut mycorrhiza, which comprises the following steps:
1) separating flesh from the boletus edulis sporocarp by adopting a tissue separation method, inoculating the flesh on a PDA (personal digital Assistant) plate culture medium, placing the culture medium in an incubator to culture and sprout hyphae at the temperature of 22-24 ℃, and transferring the hyphae to a new PDA plate culture medium again for purification culture to obtain pure hyphae;
2) after the hyphae grow over the culture dish, selecting the hyphae on an improved MMN liquid culture medium, and placing the hyphae in a shaking table for liquid fermentation culture for 12 days;
3) the greenhouse is improved, an air filtering system is added, ventilation and light transmission are kept, no mixed bacteria pollution exists in the air, and the temperature in the greenhouse is controlled to be 8-28 ℃;
4) preparing a Chinese chestnut seedling liquid nutrient solution and carrying out sterilization treatment, wherein each liter of water contains 920mg of calcium nitrate, 600 mg of potassium nitrate, 120 mg of ammonium dihydrogen phosphate and 495 mg of magnesium sulfate;
5) sterilizing and disinfecting the surfaces of chestnut seedling seeds by using a diluted potassium permanganate solution;
6) after facilities and equipment in the greenhouse are sterilized by ozone, placing the seeds sterilized in the step 5) into a seedling raising device, and injecting prepared Chinese chestnut seedling liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
7) keeping the nutrient solution in the seedling raising device to circularly flow, simultaneously filtering and sterilizing the nutrient solution flowing out of the seedling raising device by adopting a filtering method, and injecting the sterilized nutrient solution into the seedling raising device;
during the culture period, the height of the liquid level is controlled every day to ensure that the root system of the newly germinated Chinese chestnut seedlings can contact the air every day;
8) measuring the pH value of the nutrient solution by using a pH meter, and adjusting the pH value to 5.5 by using diluted hydrochloric acid or sodium hydroxide;
9) after 3 lateral roots grow out of the Chinese chestnut seedlings, closing a filtering and sterilizing system, and replacing the nutrient solution with boletus edulis liquid diluted to 1200 times;
after the replacement of the boletus edulis liquid solution, the liquid level is controlled 1 time every 3 hours, the root system of the newly germinated chestnut seedling can be contacted with the air every day, and the root is preferably kept moist; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
10) after culturing for 15 days, obtaining the boletus edulis mycorrhiza of the Chinese chestnut.
The mycorrhiza synthesis rate of the embodiment reaches 96%, the preparation method is simple, practical and efficient, the production cost is low, the influence of seasons is avoided, and the economic benefit is remarkable.
Example 4A mycorrhiza synthetic culture method of Tuber indicum and Quercus alpina
This example is a mycorrhiza synthesis culture method of tuber indicum and quercus acutissima, which is performed according to the following steps:
1) cleaning mature undamaged fruit bodies of tuber indicum, carrying out surface disinfection by using diluted alcohol, washing for 3 times by using sterile water, and adding 1000 times of sterile water into a disinfected juicer to prepare tuber indicum spore suspension;
2) the greenhouse is improved, an air filtering system is added, ventilation and light transmission are kept, no mixed bacteria pollution exists in the air, and the temperature in the greenhouse is controlled to be 8-28 ℃;
3) preparing quercus semicarpifolia seedling liquid nutrient solution and carrying out sterilization treatment, wherein each liter of water contains 900mg of calcium nitrate, 550 mg of potassium nitrate, 110 mg of ammonium dihydrogen phosphate, 490 mg of magnesium sulfate, 20mg of zinc sulfate, 10 mg of copper sulfate and 3mg of ammonium molybdate;
4) sterilizing and disinfecting the surfaces of quercus semicarpifolia seeds by using a diluted potassium permanganate solution;
5) sterilizing facilities and equipment in the greenhouse by using ozone, placing the disinfected seeds obtained in the step 4) into a seedling raising device, and injecting prepared quercus semicarpifolia seedling liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
6) keeping the nutrient solution in the seedling raising device to circularly flow, simultaneously adopting an overflowing type ultraviolet sterilizer to sterilize the nutrient solution flowing out of the seedling raising device, and injecting the sterilized nutrient solution into the seedling raising device;
during the culture period, the height of the liquid level is controlled every day to ensure that the root system of the newly germinated quercus semicarpifolia seedling can contact the air every day;
7) measuring the pH value of the nutrient solution by using a pH meter, and adjusting the pH value to 7.0-7.5 by using diluted hydrochloric acid or sodium hydroxide;
8) after 3 lateral roots grow out of the quercus semicarpifolia seedlings, closing a filtering and sterilizing system, and replacing the nutrient solution with an Indian truffle spore suspension diluted by 1000 times;
after replacing with the India truffle spore suspension, controlling the height of the liquid level 1 time every 8 hours to ensure that the root system of the newly germinated quercus semicarpifolia seedling can contact with the air every day, and preferably keeping the root wet; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
9) after culturing for 3 months, obtaining quercus acutissima roots of Indian truffles.
The mycorrhiza synthesis rate of the embodiment reaches 97%, the preparation method is simple, practical and efficient, the production cost is low, the influence of seasons is avoided, and the economic benefit is remarkable.
The embodiments 1-4 are only preferred embodiments of the present invention, but not limiting the present invention in other forms, and any person skilled in the art may make modifications or changes to the equivalent embodiments using the above technical teaching. However, simple modifications, equivalent changes and modifications of the above embodiments may be made without departing from the technical spirit of the claims of the present invention, and the scope of the claims of the present invention may be protected.
Claims (4)
1. A mycorrhiza synthesis culture method is characterized by comprising the following steps in sequence:
1-1) separating mushroom flesh from a delicious bolete fruiting body by adopting a tissue separation method, inoculating the mushroom flesh on a PDA (personal digital Assistant) plate culture medium, placing the culture medium in an incubator to culture and sprout hyphae at the temperature of 22-24 ℃, and transferring the hyphae to a new PDA plate culture medium again for purification culture to obtain pure hyphae;
1-2) after the culture dish is full of hyphae, picking the hyphae on an improved MMN liquid culture medium, and placing the hyphae in a shaking table for liquid fermentation culture for one month;
1-3) transforming the greenhouse, adding an air filtering system, keeping ventilation and light transmission, keeping no mixed bacteria pollution in the air, and controlling the temperature in the greenhouse to be 8-28 ℃;
1-4) preparing a Yunnan pine liquid nutrient solution and carrying out sterilization treatment, wherein each 100L of water contains 51g of urea, 31g of monopotassium phosphate, 10g of calcium sulfate, 5 g of magnesium sulfate, 0.01g of zinc sulfate, 0.03g of ferric sulfate, 0.01g of copper sulfate, 0.03g of manganese sulfate and 0.02g of boric acid powder;
1-5) sterilizing and disinfecting the surface of Yunnan pine seeds by using a diluted potassium permanganate solution;
1-6) sterilizing facilities in the greenhouse by ozone, placing the seeds sterilized in the step 1-5) in a seedling raising device, and injecting prepared Yunnan pine liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
1-7) keeping the nutrient solution in the seedling raising device to circularly flow, simultaneously sterilizing the nutrient solution flowing out of the seedling raising device by using an ultraviolet overflowing type sterilization system, and injecting the sterilized nutrient solution into the seedling raising device;
during the culture period, the height of the liquid level is controlled every day to ensure that the newly germinated Yunnan pine root system can contact the air every day;
1-8) measuring the pH value of the nutrient solution by using a pH measuring instrument, and adjusting the pH value to 6.0 by using diluted hydrochloric acid or sodium hydroxide;
1-9) closing the overflowing type sterilization system after 3 lateral roots of Yunnan pine grow out, and replacing the Yunnan pine liquid nutrient solution with bolete liquid diluted by 800 times;
during the culture period of the boletus liquid, the height of the liquid level is controlled every day, and the root system of the newly germinated Yunnan pine can contact the air every day; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
1-10) culturing for 2 months under the illumination condition to obtain boletus edulis mycorrhiza.
2. The mycorrhiza synthesis culture method according to claim 1, which is sequentially carried out according to the following steps:
2-1) separating spores from thelephora ganbajun sporocarp by adopting a spore separation method, inoculating the spores on a PDA (personal digital assistant) plate culture medium, placing the spores in an incubator to culture at 22-24 ℃ to sprout hyphae, transferring the hyphae to a new PDA plate culture medium again to carry out purification culture, and obtaining pure hyphae;
2-2) after the culture dish is full of hyphae, picking the hyphae on an MMN liquid culture medium, and placing the hyphae in a shaking table for liquid fermentation culture for one month;
2-3) modifying the glass greenhouse, adding a fresh air system capable of isolating bacteria, keeping ventilation and light transmission, and controlling the temperature in the glass greenhouse to be 8-28 ℃, wherein the air is free from foreign bacteria pollution;
2-4) preparing a cryptomeria fortunei liquid nutrient solution and carrying out sterilization treatment, wherein each 100L of water contains 11g of sodium nitrate, 71 g of calcium superphosphate, 25 g of ammonium sulfate, 35 g of potassium sulfate, 42 g of magnesium sulfate, 2.4 g of ferrous sulfate, 0.35 g of boric acid powder, 0.25 g of manganese sulfate and 0.2g of trace elements;
2-5) sterilizing the surface of the Yunnan oil fir seeds by using 75 percent alcohol;
2-6) stopping the propagation of fungi on the ground in the glass greenhouse by paving lime and ground cloth, carrying out spray sterilization on facilities and equipment by using 75% alcohol, placing the sterilized seeds obtained in the step 2-5) in a glass container, and injecting a taxus yunnanensis liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
2-7) keeping the circulation flow of the Yunnan fir liquid nutrient solution in the glass container, filtering and sterilizing the nutrient solution flowing out of the container by a filtering method, and injecting the sterilized nutrient solution into the glass container;
during the culture period, the liquid level of the nutrient solution is controlled to ensure that the newly germinated oil fir root system can contact the air every day;
2-8) measuring the pH value of the nutrient solution by using a pH measuring instrument, and adjusting the pH value to 6.0 by using diluted hydrochloric acid or sodium hydroxide;
2-9) closing the filtering and sterilizing system after 2 lateral roots of the Yunnan oil fir grow out, and replacing the nutrient solution with thejoram bacterial solution diluted to 1000 times;
after the rhizopus chinensis liquid is replaced by the rhizopus chinensis liquid, the height of the liquid level is controlled, and the root system of the newly germinated cryptomeria yunnanensis can be ensured to contact the air every day; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
2-10) culturing for 2 months to obtain the dry pasteurella mycorrhiza of the Yunnan oil fir.
3. The mycorrhiza synthesis culture method according to claim 1, which is sequentially carried out according to the following steps:
3-1) separating mushroom meat from the delicious bolete sporocarp by adopting a tissue separation method, inoculating the mushroom meat on a PDA (potato dextrose agar) plate culture medium, placing the mushroom meat in an incubator to culture and sprout hyphae at the temperature of 22-24 ℃, and transferring the hyphae to a new PDA plate culture medium again for purification culture to obtain pure hyphae;
3-2) after the culture dish is full of hyphae, picking the hyphae on an improved MMN liquid culture medium, and placing the hyphae in a shaking table for liquid fermentation culture for 12 days;
3-3) transforming the greenhouse, adding an air filtering system, keeping ventilation and light transmission, keeping no mixed bacteria pollution in the air, and controlling the temperature in the greenhouse to be 8-28 ℃;
3-4) preparing a Chinese chestnut seedling liquid nutrient solution and carrying out sterilization treatment, wherein each liter of water contains 920mg of calcium nitrate, 600 mg of potassium nitrate, 120 mg of ammonium dihydrogen phosphate and 495 mg of magnesium sulfate;
3-5) sterilizing the surface of the chestnut seedling seeds by using the diluted potassium permanganate solution;
3-6) sterilizing facilities in the greenhouse by ozone, placing the seeds sterilized in the step 3-5) in a seedling raising device, and injecting prepared liquid nutrient solution of the chestnut seedlings, wherein the liquid is preferably used for submerging the seeds;
3-7) keeping the nutrient solution in the seedling raising device to circularly flow, simultaneously filtering and sterilizing the nutrient solution flowing out of the seedling raising device by adopting a filtering method, and injecting the sterilized nutrient solution into the seedling raising device;
during the culture period, the height of the liquid level is controlled every day to ensure that the root system of the newly germinated Chinese chestnut seedlings can contact the air every day;
3-8) measuring the pH value of the nutrient solution by using a pH measuring instrument, and adjusting the pH value to 5.5 by using diluted hydrochloric acid or sodium hydroxide;
3-9) closing the filtering and sterilizing system after 3 lateral roots grow out of the Chinese chestnut seedlings, and replacing the nutrient solution with boletus edulis bacterial solution diluted to 1200 times;
after the replacement of the boletus edulis liquid solution, the liquid level is controlled 1 time every 3 hours, the root system of the newly germinated chestnut seedling can be contacted with the air every day, and the root is preferably kept moist; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
3-10) culturing for 15 days to obtain the boletus edulis mycorrhiza of the Chinese chestnut.
4. The mycorrhiza synthesis culture method according to claim 1, which is sequentially carried out according to the following steps:
4-1) washing mature and undamaged fruit bodies of the tuber indicum, disinfecting the surfaces of the fruit bodies by using diluted alcohol, washing the fruit bodies for 3 times by using sterile water, and adding 1000 times of sterile water into the fruit bodies of the tuber indicum by using a disinfected juicer to prepare a spore suspension of the tuber indicum;
4-2) transforming the greenhouse, adding an air filtering system, keeping ventilation and light transmission, keeping no mixed bacteria pollution in the air, and controlling the temperature in the greenhouse to be 8-28 ℃;
4-3) preparing a quercus semicarpifolia seedling liquid nutrient solution and carrying out sterilization treatment, wherein each liter of water contains 900mg of calcium nitrate, 550 mg of potassium nitrate, 110 mg of ammonium dihydrogen phosphate, 490 mg of magnesium sulfate, 20mg of zinc sulfate, 10 mg of copper sulfate and 3mg of ammonium molybdate;
4-4) sterilizing the surface of the quercus semicarpifolia seedling seeds by using the diluted potassium permanganate solution;
4-5) sterilizing facilities in the greenhouse by ozone, placing the seeds sterilized in the step 4-4) in a seedling raising device, and injecting prepared quercus semicarpifolia seedling liquid nutrient solution, wherein the liquid is preferably used for submerging the seeds;
4-6) keeping the nutrient solution in the seedling raising device to circularly flow, simultaneously adopting an over-flow type ultraviolet sterilizer to sterilize the nutrient solution flowing out of the seedling raising device, and injecting the sterilized nutrient solution into the seedling raising device;
during the culture period, the height of the liquid level is controlled every day to ensure that the root system of the newly germinated quercus semicarpifolia seedling can contact the air every day;
4-7) measuring the pH value of the nutrient solution by using a pH measuring instrument, and adjusting the pH value to 7.0-7.5 by using diluted hydrochloric acid or sodium hydroxide;
4-8) closing the filtration and sterilization system after the quercus semicarpifolia seedlings grow out of 3 lateral roots, and replacing the nutrient solution with an Indian truffle spore suspension diluted by 1000 times;
after replacing with the India truffle spore suspension, controlling the height of the liquid level 1 time every 8 hours to ensure that the root system of the newly germinated quercus semicarpifolia seedling can contact with the air every day, and preferably keeping the root wet; the sunshade net is opened every morning or evening to ensure sufficient sunlight;
4-9) culturing for 3 months to obtain the root of Quercus alpina Achillea India mycorrhiza.
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| CN105075632A (en) * | 2015-09-01 | 2015-11-25 | 大理苍山植物园生物科技有限公司 | Rhododendron decorum aseptic seedling mycorrhization technology |
| CN105917972A (en) * | 2016-07-04 | 2016-09-07 | 中国科学院昆明植物研究所 | Synthesis method of ectotrophic mycorrhiza |
| CN106576910B (en) * | 2016-12-14 | 2020-04-21 | 山西省农业科学院食用菌研究所 | Culture method of in-vitro sterile host plant-mycorrhizal edible fungus symbiotic seedling |
| CN107083335B (en) * | 2017-04-25 | 2020-05-26 | 鲁东大学 | Method for rapidly mycorrhizating DSE fungi and blueberry tissue culture seedlings |
| CN107711290B (en) * | 2017-09-28 | 2020-07-24 | 山西省农业科学院食用菌研究所 | Culture medium for mycorrhizal edible fungus symbiotic seedling and synchronous culture method thereof |
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