CN112273228B - Cultivation method of Yunnan pine mycorrhizal seedlings - Google Patents
Cultivation method of Yunnan pine mycorrhizal seedlings Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/001—Culture apparatus for tissue culture
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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
- A01G17/00—Cultivation of hops, vines, fruit trees, or like trees
- A01G17/005—Cultivation methods
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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
- A01G18/20—Culture media, e.g. compost
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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
- A01G18/40—Cultivation of spawn
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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
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/10—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material
- A01G24/12—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material containing soil minerals
- A01G24/15—Calcined rock, e.g. perlite, vermiculite or clay aggregates
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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
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/20—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material
- A01G24/28—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material containing peat, moss or sphagnum
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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
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/50—Growth substrates; Culture media; Apparatus or methods therefor contained within a flexible envelope
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/10—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in agriculture
- Y02A40/22—Improving land use; Improving water use or availability; Controlling erosion
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/40—Afforestation or reforestation
Abstract
The invention provides a cultivation method of Yunnan pine mycorrhizal seedlings, which comprises the following steps: (1) preparing strains, (2) cultivating mycorrhizal seedlings, and (3) transplanting seedlings. The cultivation method provided by the invention can help Yunnan pine seedlings to improve the stress resistance, greatly improve the survival rate of the seedlings, improve the quality of artificial forests, and enhance the stability of the forests, so that the purposes of increasing the wood reserves, giving full play to ecological benefits and creating a beautiful environment are achieved, and a foundation is laid for high-efficiency directional cultivation of Yunnan pine mycorrhiza and industrial cultivation of mycorrhizal edible fungi.
Description
Technical Field
The invention relates to the technical field of agricultural planting, in particular to a cultivation method of Yunnan pine mycorrhizal seedlings.
Background
Evergreen conifer trees of Pinaceae Pinus of Pinaceae of Pinus of Yunnan Pinus (Pinus yunnanensis) are pioneer tree species and important material tree species for afforestation of barren mountains in southwest regions, and have high economic value and ecological benefit. However, local masses fell and utilize the trees in a mode of cutting large and leaving small, cutting straight and leaving bent for a long time, so that the quality of the Yunnan pine forest is reduced, the proportion of the ' bent and twisted pine and the ' ground pine ' is continuously increased, the development of high-quality and high-efficiency artificial forests of the Yunnan pine is seriously restricted, the utilization of wood is mainly achieved by cutting the natural Yunnan pine forest, the quality of the whole Yunnan pine forest stand is further influenced, and large-area low-quality and low-efficiency forests are further generated.
Abundant Ectomycorrhizal Edible Fungi (EEF) are stored in Yunnan pine forest. The mycorrhizal edible fungi are not only important agricultural and sideline products in farmer markets in various places of Yunnan, but also one of important commodities for earning foreign exchange through natural resource exports in Yunnan province. At present, a mycorrhizal edible fungi cultivation mode which can not be artificially cultivated but can really form a mutual and beneficial relationship with trees becomes a research direction, the cultivation mode is a 'seedling cultivation and forestation-mycorrhizal edible fungi mode', namely, a tree species and precious edible mycorrhizal fungi species which can be symbiotic with the tree species are selected before forestation, the mycorrhiza is artificially synthesized, the mode is changed from single forestation as a main mode to seedling cultivation and precious edible mycorrhizal fungi as a main mode, the economic income is increased, and the advantages of the mycorrhizal symbiotic relationship are fully exerted. At present, mycorrhizal edible fungi become a new pet in agriculture and forestry, and not only have unique edible and medicinal values, but also can promote the growth of trees, improve the yield and enhance the stress resistance of forest trees. Unfortunately, it is not possible to cultivate completely artificially, but only by using mycorrhiza technology, relying on the roots of the host plant for "mycorrhiza synthesis".
The phenomenon of squat seedling is often found in the Yunnan pine seedling raising process, and is attributed to poor drought resistance and disease resistance. The cultivation of mycorrhizal seedlings can help Yunnan pine seedlings to improve the stress resistance, the survival rate of the seedlings is greatly improved, the quality of artificial forests is improved, the stability of the forests is enhanced, the purposes of increasing wood reserves, giving full play to ecological benefits and creating a beautiful environment are further achieved, and a foundation is laid for the efficient and directional cultivation of mycorrhizal fungi of Yunnan pine and the industrialized cultivation of mycorrhizal edible fungi.
Disclosure of Invention
The invention aims to provide a cultivation method of Yunnan pine mycorrhizal seedlings, so as to improve the survival rate of the Yunnan pine seedlings and simultaneously improve the yield of mycorrhizal edible fungi.
A cultivation method of Yunnan pine mycorrhizal seedlings comprises the following steps:
(1) strain preparation
Collecting fresh sporophore of Cephalosporium nipponensis, selecting one tissue on the fresh sporophore of Cephalosporium nipponensis as culture tissue block, selecting it into test tube slant of comprehensive PDA culture medium by using inoculating needle, placing in 25 deg.C incubator and dark-culturing for 30d, and continuously culturing for 40d after the grown hypha is transferred to MMN culture medium to obtain spare bacterial block;
wherein, the comprehensive PDA culture medium is as follows: in 1L, the method comprises the following steps: 200g of potatoes, 20g of glucose, 3g of monopotassium phosphate, 1.5g of magnesium sulfate, 10mg of vitamins and 18g of agar, and sterilizing for 20min at 121 ℃;
the MMN culture medium is as follows: in 1L, comprises: 15.0g of agar, 0.025g of NaCl, 10g of glucose, 0.20g of citric acid and KH 2 PO 4 0.50g, malt extract powder 3g, vitaminBiotin B10.1mg, CaCl 2 ·2H 2 O 0.05g，(NH 4 ) 2 HPO 4 0.25g of FeCl with the mass percent concentration of 1 percent 3 1.2mL，MgSO 4 ·7H 2 O 0.15g，pH 5.7±0.1；
(2) Cultivation of mycorrhizal seedlings
Transferring the spare Yunnan pine seeds which are germinated in advance into the tissue culture bottles under an aseptic condition, and inoculating spare bacterium blocks around roots in each tissue culture bottle after the seeds germinate and grow young roots; sealing with air-permeable bacteria-isolating membrane, culturing in light culture chamber for 6 months, and wrapping the culture bottle with tinfoil paper to prevent the strain and root system from directly exposing to light;
the tissue culture bottle is internally provided with an improved MMN solid culture medium, and the improved MMN solid culture medium is as follows:
in 1L, comprises: 5g of yeast extract, 3g of malt extract powder, 16.0g of agar, 10g of glucose, 2g of peptone, 0.10g of citric acid, 0.025g of NaCl and KH 2 PO 4 0.50g，CaCl 2 ·2H 2 0.5g of FeCl with the mass percent concentration of 1 percent 3 1.2mL，MgSO 4 ·7H 2 O 0.15g，(NH 4 ) 2 HPO 4 0.25g, vitamin B10.1mg, IBA 1.0mg, NAA 0.10mg, active carbon 0.50g, pH 5.5-6.0;
(3) transplanting of seedlings
And (3) filling the seedling culture substrate subjected to irradiation sterilization into a standby seedling culture bag, and transplanting the mycorrhizal seedlings obtained in the step (2) into the seedling culture bag.
Further, in the method for cultivating the Yunnan pine mycorrhizal seedlings, the selection mode of the cultured tissue blocks in the step (1) is as follows: a small piece of tissue is picked up at the junction or fold of cap and stipe of fresh fruiting body of Cephalosporium nipponicum, and cut into tissue pieces of about 1cm × 1.5cm and 1.5mm in thickness to serve as a cultured tissue piece.
Further, the cultivation method of the Yunnan pine mycorrhizal seedlings further comprises the step of cleaning fresh sporocarp of the green-head fungus before selecting the culture tissue block, and specifically comprises the following steps: carefully remove the soil adhered to the pileus and stipe, and wipe with 75% alcohol cotton ball to sterilize for 2 times.
Further, according to the cultivation method of the Yunnan pine mushroom root seedling, the green head mushroom fresh fruit body is 70% -80% in maturity and free of diseases and insect pests.
Further, according to the cultivation method of the Yunnan pine mycorrhizal seedlings, in the step (2), the area of the standby mycorrhiza block which is inoculated to the periphery of the root in the tissue culture bottle is 1cm 2 The number of the inoculated bacteria is 3.
Further, in the cultivation method of Yunnan pine mycorrhizal seedlings, the tissue culture bottle filled with the improved MMN solid culture medium in the step (2) needs to be sterilized at high temperature and high pressure for 20min before use.
Further, in the method for cultivating Yunnan pine mycorrhizal seedlings as described above, the seedling raising bag in step (3) is sterilized with 10% hydrogen peroxide solution before use, then washed with tap water, and finally washed with sterile water, dried in the air and kept ready for use.
Further, according to the cultivation method of the Yunnan pine mycorrhizal seedlings, the filter paper for preventing the substrate from leaking is laid at the bottom of the seedling raising bag in the step (3).
Further, in the method for cultivating the Yunnan pine mycorrhizal seedlings, the seedling substrate in the step (3) is a substrate prepared from the following components in a volume ratio of 1: 1 mixture of vermiculite and peat soil.
Has the advantages that:
according to the cultivation method of the Yunnan pine mycorrhizal seedlings, strain preparation and mycorrhizal seedling cultivation under the aseptic condition are comprehensively utilized, successful inoculation and aseptic matrix transplanting are combined, the single infection rate of edible mycorrhizal fungi, namely the green head fungi is remarkably improved, mycorrhiza synthesis of the Yunnan pine mycorrhizal seedlings and the green head fungi is promoted, and a foundation is laid for the next formation of green head fungi sporocarp. Meanwhile, after mycorrhiza is formed by the Yunnan pine seedlings and the cephalosporium, the growth index of the Yunnan pine seedlings can be obviously improved, and the antioxidant activity of the Yunnan pine seedlings is enhanced, so that excellent Yunnan pine seedlings are cultured. In addition, the culture medium of the cultivation method is light in weight, convenient to transport and beneficial to mountain-climbing afforestation.
Drawings
FIG. 1 is a mycorrhizal diagram formed by the root system of Yunnan pine seedlings cultivated by the method of the invention.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention are described clearly and completely below, and it is obvious that the described embodiments are some, not all embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
1 study materials
1.1 test materials
The Cephalotaxus fortunei fruiting body is derived from fresh edible fungi newly collected from Yunnan Pinus pure forest. The Yunnan pine seeds come from a first-generation Yunnan pine seed garden in the Kunming tree garden. Seeds were treated with 10% H on a sterile bench 2 O 2 Treating for 10min, washing with sterile water for 5 times, air drying, and culturing in sterile culture dish with diameter of 9cm and 1 layer of filter paper. The culture environment is at 25 + -1 deg.C, and the light is 14 hr and dark is 10 hr per day. After the culture for 21d, when the radicle grows to 1 cm-2 cm, the product is ready for use.
1.2 culture Medium
Modified MMN medium (1L): 5g of yeast extract, 3g of malt extract powder, 16.0g of agar, 10g of glucose, 2g of peptone, 0.10g of citric acid, 0.025g of NaCl and KH 2 PO 4 0.50g，CaCl 2 ·2H 2 O 0.5g，FeCl 3 (1% solution) 1.2mL, MgSO 4 ·7H 2 O 0.15g，(NH 4 ) 2 HPO 4 0.25g, vitamin B10.1mg, IBA 1.0mg, NAA 0.10mg, active carbon 0.50g, pH 5.5-6.0.
Standard MMN medium (1L): 15.0g of agar, 0.025g of NaCl, 10g of glucose, 0.20g of citric acid and KH 2 PO 4 0.50g, malt extract powder 3g, vitamin B10.1mg, CaCl 2 ·2H 2 O 0.05g，(NH 4 ) 2 HPO 4 0.25g，FeCl 3 (1% solution) 1.2mL, MgSO 4 ·7H 2 O 0.15g，pH 5.7±0.1。
The improved MMN culture medium is added with growth hormones such as IBA and NAA for promoting plant rooting, which is helpful for a target host to root in the culture medium and promote mycorrhiza synthesis. The addition of the active carbon can provide a dark environment for hypha growth, prevent browning, improve the content of soluble protein and total sugar in the culture medium and promote hypha growth and infection. The yeast extract and the peptone are added to provide rich nitrogen sources for the growth of mycorrhizal fungi and plants and promote the growth of the mycorrhizal fungi and the plants. Meanwhile, experiments show that the formula of the improved MMN culture medium is most suitable for mycorrhiza synthesis.
2 research methods
2.1 Strain preparation
Collecting 70% -80% mature fresh fruiting body of Trichophyton mentagrophytes (Russulvirescens) without diseases and insect pests, carefully removing the adhered soil on pileus and stipe, wiping with 75% alcohol cotton ball for sterilization, picking a small tissue at the junction or fold of pileus and stipe, cutting into tissue blocks with size of about 1cm × 1.5cm and thickness of 1.5mm, picking into PDA culture medium test tube slant with inoculating needle, placing in 25 deg.C incubator in dark for culturing for 30d, and transferring grown hypha to standard MMN culture medium for culturing for 40 d. Under a clean bench, a sterilization punch with the diameter of 5mm is used for punching holes on the edges of the bacterial colonies for later use.
2.2 cultivation of mycorrhizal seedlings
The improved MMN culture medium is filled into a 500mL tissue culture bottle, sterilized at high temperature and high pressure for 20min, and moved to an ultra-clean workbench for standby. In a superclean workbench, moving the standby Yunnan pine seeds which germinate in advance into the tissue culture bottles under the aseptic condition, and after the seeds germinate and grow young roots, accessing the standby area of 1cm around the roots in each tissue culture bottle 2 The 3 fungus blocks are sealed (ventilated), then the culture is placed in a light culture room for 6 months, and the tissue culture bottle is wrapped by tinfoil paper to prevent the strains and the root systems from being directly exposed to light.
2.3 transplanting of seedlings
Sterilizing 12 × 13cm seedling bag with 10% hydrogen peroxide solution for 10min, washing with tap water for three times, washing with sterile water, air drying, and placing a piece of filter paper at the bottom to prevent matrix leakage. The seedling substrate is a mixture of vermiculite and peat soil (the volume ratio is 1: 1), and the mixture is irradiated and sterilized and then is put into a seedling bag for later use.
After the mycorrhizal seedlings are transplanted, watering, weeding and fertilizing are carried out, herbicide is forbidden to be used during weeding, ventilation of a nursery is guaranteed, diseases are prevented, and no fungicide is applied during seedling raising.
3 results of the experiment
TABLE 1 influence of different treatments on Yunnan pine seedling growth and root infection rate under different substrate conditions
From table 1, it can be seen that: the growth indexes (infection rate, root branch number, plant height, fresh weight and dry weight) of the Yunnan pine seedlings inoculated with the eurotium cristatum strain are obviously higher than those of the Yunnan pine seedlings not inoculated with the eurotium cristatum strain, wherein the infection rate of the ectomycorrhiza of the Yunnan pine seedlings not inoculated with the eurotium cristatum strain is 0. Moreover, the Yunnan pine seedlings inoculated with the inoculus vernalis strain are cultured in a culture medium with the volume ratio of 1: 1, the infection rate, the root branch number, the plant height, the fresh weight and the dry weight of the mixture seedling substrate of vermiculite and peat soil are the highest and are respectively 46.33 percent, 17.00 roots, 17.93cm, 0.92g and 0.21 g.
TABLE 2 influence of different treatments on physiological indices of pinus yunnanensis seedlings on the coniferous and root of pinus yunnanensis seedlings under different substrate conditions
From table 2, it can be seen that: in the three seedling substrates of the method, except MDA, the contents of SOD, POD, CAT and Pro of the inoculated Yunnan pine seedlings (needle leaves and roots) are obviously higher than those of a control, which shows that the physiological activity of the Yunnan pine seedlings can be obviously improved by the inoculation treatment of the cynanchum glaucescens, and various adverse situations can be better resisted. Wherein, the volume ratio is 1: the mixture seedling culture substrate of vermiculite and peat soil of 1 is most suitable for the culture and the exertion of physiological activity of Yunnan pine mycorrhizal seedlings.
FIG. 1 shows a large number of mycorrhizas formed by the root system of Yunnan pine seedlings cultivated by the method of the present invention, which proves that the method can cultivate a large number of mycorrhiza seedlings.
Finally, it should be noted that: the above examples are only intended to illustrate the technical solution of the present invention, and not to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, it should be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions of the embodiments of the present invention.
Claims (8)
1. A cultivation method of Yunnan pine mycorrhizal seedlings is characterized by comprising the following steps:
(1) strain preparation
Collecting fresh sporophore of Cephalosporium nipponensis, selecting one tissue on the fresh sporophore of Cephalosporium nipponensis as culture tissue block, selecting it into test tube slant of comprehensive PDA culture medium by using inoculating needle, placing in 25 deg.C incubator and dark-culturing for 30d, and continuously culturing for 40d after the grown hypha is transferred to MMN culture medium to obtain spare bacterial block;
wherein, the comprehensive PDA culture medium is as follows: in 1L, the method comprises the following steps: 200g of potato, 20g of glucose, 3g of monopotassium phosphate, 1.5g of magnesium sulfate, 10mg of vitamin and 18g of agar, and sterilizing for 20min at 121 ℃;
the MMN culture medium is: in 1L, the method comprises the following steps: 15.0g of agar, 0.025g of NaCl, 10g of glucose, 0.20g of citric acid and KH 2 PO 4 0.50g, malt extract powder 3g, vitamin B 1 0.1mg，CaCl 2 ·2H 2 O 0.05g，(NH 4 ) 2 HPO 4 0.25g of FeCl with the mass percent concentration of 1 percent 3 1.2 mL，MgSO 4 ·7H 2 O 0.15g，pH 5.7±0.1；
(2) Cultivation of mycorrhizal seedlings
Transferring spare Yunnan pine seeds which germinate in advance into the tissue culture bottles under an aseptic condition, and inoculating spare bacterium blocks around roots in each tissue culture bottle after the seeds germinate and grow into young roots; sealing with air-permeable bacteria-isolating membrane, culturing in light culture chamber for 6 months, and wrapping the culture bottle with tinfoil paper to prevent the strain and root system from directly exposing to light;
the tissue culture bottle is internally provided with an improved MMN solid culture medium, and the improved MMN solid culture medium is as follows:
in 1L, the method comprises the following steps: 5g of yeast extract, 3g of malt extract powder, 16.0g of agar, 10g of glucose, 2g of peptone, 0.10g of citric acid, 0.025g of NaCl and KH 2 PO 4 0.50g，CaCl 2 ·2H 2 0.5g of FeCl with the mass percent concentration of 1 percent 3 1.2 mL，MgSO 4 ·7H 2 O 0.15g，(NH 4 ) 2 HPO 4 0.25g, vitamin B 1 0.1mg, IBA 1.0mg, NAA 0.10mg, active carbon 0.50g, pH 5.5-6.0;
(3) transplanting of seedlings
Placing the seedling substrate subjected to irradiation sterilization into a standby seedling bag, wherein the seedling substrate is prepared from the following components in a volume ratio of 1: 1, transplanting the mycorrhizal seedlings obtained in the step (2) into the seedling raising bag.
2. The cultivation method of Yunnan pine mycorrhizal seedlings according to claim 1, characterized in that the selection mode of the cultivation tissue block in the step (1) is as follows: a small piece of tissue is picked up at the junction or fold of cap and stipe of fresh fruiting body of Cephalosporium nipponicum, and cut into tissue blocks of about 1cm × 1.5cm and 1.5mm thickness as culture tissue blocks.
3. A cultivation method of Yunnan pine mycorrhizal seedlings according to claim 1, which is characterized by further comprising a cleaning step of fresh fruiting bodies of the green-headed fungus before selecting the cultivation tissue blocks, specifically comprising the following steps: carefully remove the soil adhered to the pileus and stipe, and wipe with 75% alcohol cotton ball to sterilize for 2 times.
4. The cultivation method of Yunnan pine mycorrhizal seedlings according to claim 1, wherein the fresh fruit body of the green-head mushroom is 70% -80% mature without plant diseases and insect pests.
5. The method for cultivating Yunnan pine mycorrhizal seedlings according to claim 1, wherein in the step (2), the area of the spare mycorrhiza block which is inoculated to the periphery of the root in the tissue culture bottle is 1cm 2 The number of the inoculated bacteria is 3.
6. The cultivation method of Yunnan pine mycorrhizal seedlings according to claim 1, wherein the tissue culture bottle filled with the improved MMN solid medium in the step (2) is sterilized at high temperature and high pressure for 20min before use.
7. The cultivation method of Yunnan pine mycorrhizal seedlings according to claim 1, characterized in that, before the bag for raising seedlings in step (3) is used, the bag for raising seedlings is sterilized by 10% hydrogen peroxide solution, then washed by tap water, finally washed by sterile water, dried by air and prepared for use.
8. The cultivation method of Yunnan pine mycorrhizal seedlings according to claim 1, wherein the filter paper for preventing the substrate from leaking is laid on the bottom of the seedling raising bag in the step (3).
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| CN116904325B (en) * | 2023-09-08 | 2023-12-15 | 哈尔滨市农业科学院 | High-temperature-resistant coralloid hericium erinaceus strain with high dextran yield and breeding and cultivation method |
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| CN108432547A (en) * | 2018-03-14 | 2018-08-24 | 云南上智科技有限公司 | A kind of mycorhiza synthesis cultural method |
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| WO2013148328A1 (en) * | 2012-03-26 | 2013-10-03 | Pronutria, Inc. | Nutritive proteins and methods |
| CN106922393A (en) * | 2017-03-11 | 2017-07-07 | 云南绿辰生物科技开发有限公司 | A kind of synthetic method of truffle mycorrhiza fungi seeding |
| CN107711290A (en) * | 2017-09-28 | 2018-02-23 | 山西省农业科学院食用菌研究所 | The culture medium and its synchronous cultural method of mycorhiza edible mushroom symbiosis seedling |
| CN108432547A (en) * | 2018-03-14 | 2018-08-24 | 云南上智科技有限公司 | A kind of mycorhiza synthesis cultural method |
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