CN109006189A - A method of utilizing monokaryon Protoplast Technique breeding needle mushroom excellent variety - Google Patents
A method of utilizing monokaryon Protoplast Technique breeding needle mushroom excellent variety Download PDFInfo
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- CN109006189A CN109006189A CN201810878454.7A CN201810878454A CN109006189A CN 109006189 A CN109006189 A CN 109006189A CN 201810878454 A CN201810878454 A CN 201810878454A CN 109006189 A CN109006189 A CN 109006189A
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- protoplast
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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
- A01G18/00—Cultivation of mushrooms
-
- 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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- Mycology (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
This application involves Flammulina velutipes Protoplasts separation, monokaryon protoplast identification, cultivation fruiting, be separately cultured with excellent variety screening the methods of, using this method obtain kind inheritance stability, heredity and breeding on be of great significance.
Description
Technical field
This application involves Flammulina velutipes Protoplasts separation, monokaryon protoplast identification, cultivation fruiting, be separately cultured with it is excellent
Method for screening varieties.
Background technique
Acupuncture needle massee fruiting bodies are grown thickly, and base portion has meristematic capacity, and mature acupuncture needle massee fruiting bodies are by cap, lamella, stem three
Part forms.To be spherical to hemispherical when cap is immature, rear to be gradually spread out again as flat umbrella, surface is glossy, sticks when wet
It is sliding, it is glossy when doing, there is colloid thin skin on cap, edge is very thin, and bacteria cover diameter about 1-10cm is differed under natural conditions.Needle mushroom
It is famous one of edible mushroom of all times, integrates " natural, nutrition, health care ", the moisture content of fresh needle mushroom is about
82%-89% is rich in a large amount of protein, carbohydrate, mineral element, vitamin and crude fibre, and fat content is lower, is one
The rare high nutrition good merchantable brand of kind.
The growth and development of needle mushroom needs carbon nitrogen source, mineral element, vitamin, suitable temperature, humidity, air and light
According to.It in the prior art, include: wild domesticated breeding, crossbreeding and mutation breeding for the selection of needle mushroom.But
Above-mentioned breeding method can not all solve the problems, such as needle mushroom Genomic instability in the prior art.
Summary of the invention
The application's is designed to provide a kind of method using monokaryon Protoplast Technique breeding needle mushroom excellent variety,
By the protoplast electrofusion of this method, the identification of monokaryon protoplast, cultivation fruiting, it is separately cultured and is screened with excellent variety, with
Obtain the excellent Varieties in Flammulina velutipes of kind inheritance stability.
The application uses following technical scheme:
1. protoplast electrofusion method
1.1 Mycelium culture
Strain is dicaryon, and culture medium is 5 ~ 6g of yeast powder, 18 ~ 20g of glucose, water 1000ml；The culture medium of preparation is packed into
In 300ml triangular flask, loading amount 50ml sterilizes 30 minutes at 121 DEG C.The strain of 0.5cm × 0.5cm size is taken to be put into training fastly
It supports in base, is cultivated at 22 ~ 25 DEG C, incubation time is 8 ~ 10 days.
1.2 enzyme solution preparation methods
The enzyme for dissolving cell wall is lywallzyme (Lywallzyme), and compound concentration is 10% ~ 12%, with 0.5 ~ 0.6mol/L MgSO4
Liquid is prepared.
1.3 protoplast electrofusion methods
It takes 3~5g of mycelium of needle mushroom to be put into 50 ml triangular flasks, 10 ~ 15ml of enzyme solution is added, shakes 3 at 30~32 DEG C
~4h is filtered with sterile G2 funnel, collects filtrate in test tube, (5000 revs/min) removal enzyme solutions are centrifuged on centrifuge, heavy
It is 0.6mol/L MgSO that concentration is added in shallow lake4Sterile liquid is diluted to 1 × 103A/ml, as Protoplast suspension.
1.4 protoplast regeneration
1.4.1 regeneration culture medium production method: regeneration culture medium formula is 180 ~ 200g of potato, 100 ~ 110g of sucrose, agar
20g, water 1000ml.Peeling potatoes are cut into slices, are put into pot, water 1000ml are added, heating is boiled 30 minutes, with 4 layers of yarn
Cloth filtering, is added agar in filtrate, continues heating and boils until agar melts completely, sucrose, heating stirring is added in fusion
Melt to sucrose, be eventually adding water and be settled to 1000ml, measures culture medium 200ml and be packed into the triangular flask of 250 ml, with sealing
Film sealing, sterilization treatment 30 minutes, culture medium is poured into sterile petri dish at 121 DEG C, in culture dish culture base unit weight be 20 ~
25ml is plating medium after cooling.
1.4.2 inoculated and cultured method: taking 0.1ml Protoplast suspension to be put on plating medium, will be former with glass bar
Raw plastid smearing is dispersed in plating medium surface, then, is put into culture in 20 ~ 25 DEG C of incubator, cultivates 8-10 days, training
It after feeding primary surface grows white colony, takes single bacterium colony with inoculating tool, is put into slant medium (culture medium prescription is with 1.1)
On, it is cultivated at 20 ~ 25 DEG C, growing is Protoplast regeneration strains after mycelium.
The identification of 1.5 monokaryon protoplasts
It takes mycelium to be placed on glass slide, drips water, covered is observed under the microscope, without clamp connection on mycelia
Label is monokaryon Protoplast fusion strain.
2. cultivating fruiting method
2.1 culture medium prescriptions and sterilizing methods
Cotton seed hulls or corncob 78%, wheat bran or rice bran 30%, calcium carbonate 2%, water content 65%.Culture medium is prepared by formula rate,
It is then charged into 1100ml plastic bottle, sterilization treatment 2 hours at 121 DEG C.
2.2 inoculated and cultured methods
Strain is accessed under gnotobasis in transfer room, bacterium germination is cultivated at 20-22 DEG C, is cultivated 23-25 days, mycelial growth is full
After bottle, into management of producing mushroom.
2.3 fruiting method
The strain that mycelial growth expires bottle is moved on in cultivating chamber, cap removing is gone, digs out surface layer strain, 10 ~ 15 DEG C of temperature, air
Relative humidity 95%, 10 ~ 50 lux of intensity of illumination.Reach 14cm ~ 16cm by 22 ~ 24 days acupuncture needle massee fruiting bodies length
Harvesting.
3. organizing isolated culture method
It takes fructification to carry out tissue separation, cuts stem with scalpel and cap handover site tissue is placed in PDA culture medium,
It is cultivated at 25 DEG C, after culture 5 days, selection carries out switching culture without the vigorous strain of miscellaneous bacteria, mycelial growth, transfers oblique in PDA
On the culture medium of face, 8 ~ 10 days are cultivated at 25 DEG C, as the strain that is separately cultured of tissue.
Test yield, cap morphological feature, stem length, diameter etc. filter out excellent variety.
4. excellent variety screening technique
4.1 cultivation fruiting methods
Fruiting method is cultivated with " 2. cultivation fruiting method ".
4.2 excellent variety screening techniques
It by the test to breeding time, yield, sporophore shape feature etc., is compared with cultivar, screening yield is high
In control, it is quality also in the bacterial strain of control, as excellent variety.Using the Varieties in Flammulina velutipes inheritance stability of this method breeding,
Selection is easy.
Specific embodiment
Embodiment one
1. protoplast electrofusion method
1.1 Mycelium culture
Strain is dicaryon, and culture medium is yeast powder 5g, glucose 18g, water 1000ml；The culture medium of preparation is packed into 300ml
In triangular flask, loading amount 50ml sterilizes 30 minutes at 121 DEG C.The strain of 0.5cm × 0.5cm size is taken to be put into culture medium fastly
In, it is cultivated under 22, incubation time is 8 days.
1.2 enzyme solution preparation methods
The enzyme for dissolving cell wall is lywallzyme (Lywallzyme), and compound concentration is 10% ~ 12%, with 0.5mol/L MgSO4Liquid is matched
System.
1.3 protoplast electrofusion methods
It takes the mycelium 3g of needle mushroom to be put into 50 ml triangular flasks, enzyme solution 10ml is added, 3h is shaken at 30~32 DEG C, with nothing
The G2 funnel of bacterium filters, and collects filtrate in test tube, (5000 revs/min) removal enzyme solutions are centrifuged on centrifuge, are added in precipitating
Concentration is 0.6mol/L MgSO4Sterile liquid is diluted to 1 × 103A/ml, as Protoplast suspension.
1.4 protoplast regeneration
1.4.1 regeneration culture medium production method: regeneration culture medium formula is potato 180g, sucrose 100g, agar 20g, water
1000ml.Peeling potatoes to be cut into slices, are put into pot, water 1000ml is added, heating is boiled 30 minutes, with 4 layers of filtered through gauze,
Agar is added in filtrate, continues heating and boils until agar melts completely, sucrose is added in fusion, and heating stirring to sucrose is melted
Change, be eventually adding water and be settled to 1000ml, measures culture medium 200ml and be packed into the triangular flask of 250 ml, sealed with sealed membrane,
Sterilization treatment 30 minutes, culture medium is poured into sterile petri dish at 121 DEG C, and culture base unit weight is 20ml in culture dish, after cooling
As plating medium.
1.4.2 inoculated and cultured method: taking 0.1ml Protoplast suspension to be put on plating medium, will be former with glass bar
Raw plastid smearing is dispersed in plating medium surface, then, is put into culture in 20 DEG C of incubator, cultivates 8 days, in culture base table
It looks unfamiliar after growing white colony, takes single bacterium colony with inoculating tool, be put on slant medium (culture medium prescription is with 1.1),
It is cultivated at 20 DEG C, growing is Protoplast regeneration strains after mycelium.
The identification of 1.5 monokaryon protoplasts
It takes mycelium to be placed on glass slide, drips water, covered is observed under the microscope, without clamp connection on mycelia
Label is monokaryon Protoplast fusion strain.
2. cultivating fruiting method
2.1 culture medium prescriptions and sterilizing methods
Cotton seed hulls or corncob 78%, wheat bran or rice bran 30%, calcium carbonate 2%, water content 65%.Culture medium is prepared by formula rate,
It is then charged into 1100ml plastic bottle, sterilization treatment 2 hours at 121 DEG C.
2.2 inoculated and cultured methods
Strain is accessed under gnotobasis in transfer room, bacterium germination is cultivated at 20 DEG C, is cultivated 23 days, after mycelial growth expires bottle,
Into management of producing mushroom.
2.3 fruiting method
The strain that mycelial growth expires bottle is moved on in cultivating chamber, cap removing is gone, digs out surface layer strain, 10 DEG C of temperature, air phase
To humidity 95%, 10 lux of intensity of illumination.Reach 14cm ~ 16cm by 22 days acupuncture needle massee fruiting bodies length, can harvest.
3. organizing isolated culture method
It takes fructification to carry out tissue separation, cuts stem with scalpel and cap handover site tissue is placed in PDA culture medium,
It is cultivated at 25 DEG C, after culture 5 days, selection carries out switching culture without the vigorous strain of miscellaneous bacteria, mycelial growth, transfers oblique in PDA
On the culture medium of face, 8 days are cultivated at 25 DEG C, as the strain that is separately cultured of tissue.
Test yield, cap morphological feature, stem length, diameter etc. filter out excellent variety.
4. excellent variety screening technique
4.1 cultivation fruiting methods
Fruiting method is cultivated with " 2. cultivation fruiting method ".
4.2 excellent variety screening techniques
It by the test to breeding time, yield, sporophore shape feature etc., is compared with cultivar, screening yield is high
In control, quality also superior to the bacterial strain of control, as excellent variety.
Embodiment two
1. protoplast electrofusion method
1.1 Mycelium culture
Strain is dicaryon, and culture medium is yeast powder 6g, glucose 20g, water 1000ml；The culture medium of preparation is packed into 300ml
In triangular flask, loading amount 50ml sterilizes 30 minutes at 121 DEG C.The strain of 0.5cm × 0.5cm size is taken to be put into culture medium fastly
In, it is cultivated at 25 DEG C, incubation time is 8 ~ 10 days.
1.2 enzyme solution preparation methods
The enzyme for dissolving cell wall is lywallzyme (Lywallzyme), compound concentration 12%, with 0.6mol/L MgSO4Liquid is prepared.
1.3 protoplast electrofusion methods
It takes the mycelium 5g of needle mushroom to be put into 50 ml triangular flasks, enzyme solution 15ml is added, 4h is shaken at 30~32 DEG C, with nothing
The G2 funnel of bacterium filters, and collects filtrate in test tube, (5000 revs/min) removal enzyme solutions are centrifuged on centrifuge, are added in precipitating
Concentration is 0.6mol/L MgSO4Sterile liquid is diluted to 1 × 103A/ml, as Protoplast suspension.
1.4 protoplast regeneration
1.4.1 regeneration culture medium production method: regeneration culture medium formula is potato 200g, sucrose 110g, agar 20g, water
1000ml.Peeling potatoes to be cut into slices, are put into pot, water 1000ml is added, heating is boiled 30 minutes, with 4 layers of filtered through gauze,
Agar is added in filtrate, continues heating and boils until agar melts completely, sucrose is added in fusion, and heating stirring to sucrose is melted
Change, be eventually adding water and be settled to 1000ml, measures culture medium 200ml and be packed into the triangular flask of 250 ml, sealed with sealed membrane,
Sterilization treatment 30 minutes, culture medium is poured into sterile petri dish at 121 DEG C, and culture base unit weight is 25ml in culture dish, after cooling
As plating medium.
1.4.2 inoculated and cultured method: taking 0.1ml Protoplast suspension to be put on plating medium, will be former with glass bar
Raw plastid smearing is dispersed in plating medium surface, then, is put into culture in 25 DEG C of incubator, cultivates 10 days, in culture medium
After surface grows white colony, single bacterium colony is taken with inoculating tool, is put on slant medium (culture medium prescription is with 1.1),
It is cultivated at 25 DEG C, growing is Protoplast regeneration strains after mycelium.
The identification of 1.5 monokaryon protoplasts
It takes mycelium to be placed on glass slide, drips water, covered is observed under the microscope, without clamp connection on mycelia
Label is monokaryon Protoplast fusion strain.
2. cultivating fruiting method
2.1 culture medium prescriptions and sterilizing methods
Cotton seed hulls or corncob 78%, wheat bran or rice bran 30%, calcium carbonate 2%, water content 65%.Culture medium is prepared by formula rate,
It is then charged into 1100ml plastic bottle, sterilization treatment 2 hours at 121 DEG C.
2.2 inoculated and cultured methods
Strain is accessed under gnotobasis in transfer room, bacterium germination is cultivated at 22 DEG C, is cultivated 25 days, after mycelial growth expires bottle,
Into management of producing mushroom.
2.3 fruiting method
The strain that mycelial growth expires bottle is moved on in cultivating chamber, cap removing is gone, digs out surface layer strain, 15 DEG C of temperature, air phase
To humidity 95%, 50 lux of intensity of illumination.Reach 14cm ~ 16cm by 24 days acupuncture needle massee fruiting bodies length, can harvest.
3. organizing isolated culture method
It takes fructification to carry out tissue separation, cuts stem with scalpel and cap handover site tissue is placed in PDA culture medium,
It is cultivated at 25 DEG C, after culture 5 days, selection carries out switching culture without the vigorous strain of miscellaneous bacteria, mycelial growth, transfers oblique in PDA
On the culture medium of face, 10 days are cultivated at 25 DEG C, as the strain that is separately cultured of tissue.
Test yield, cap morphological feature, stem length, diameter etc. filter out excellent variety.
4. excellent variety screening technique
4.1 cultivation fruiting methods
Fruiting method is cultivated with " 2. cultivation fruiting method ".
4.2 excellent variety screening techniques
It by the test to breeding time, yield, sporophore shape feature etc., is compared with cultivar, screening yield is high
In control, quality also superior to the bacterial strain of control, as excellent variety.
Unquestionably, of the invention, can also have a variety of transformation and remodeling, however it is not limited to the specific reality of above embodiment
Apply example.In short, protection scope of the present invention should include those obvious transformation to those skilled in the art
Or it substitutes and retrofits.
Claims (4)
1. a kind of method using monokaryon Protoplast Technique breeding needle mushroom excellent variety, it is characterised in that by following steps group
At: protoplast electrofusion method, cultivation fruiting method, tissue isolated culture method and excellent variety screening technique.
2. the method according to claim 1 using monokaryon Protoplast Technique breeding needle mushroom excellent variety, feature
Be: the protoplast electrofusion method includes 1. Mycelium cultures；2. enzyme solution preparation method；3. protoplast electrofusion side
Method；4. protoplast regeneration；5. monokaryon protoplast is identified；Specifically:
1. Mycelium culture
Strain is dicaryon, and culture medium is 5 ~ 6g of yeast powder, 18 ~ 20g of glucose, water 1000ml；The culture medium of preparation is packed into
In 300ml triangular flask, loading amount 50ml sterilizes 30 minutes at 121 DEG C, the strain of 0.5cm × 0.5cm size is taken to be put into training fastly
It supports in base, is cultivated at 22 ~ 25 DEG C, incubation time is 8 ~ 10 days；
2. enzyme solution preparation method
The enzyme for dissolving cell wall is lywallzyme, and compound concentration is 10% ~ 12%, with 0.5 ~ 0.6mol/L MgSO4Liquid is prepared；
3. protoplast electrofusion method
It takes 3~5g of mycelium of needle mushroom to be put into 50 ml triangular flasks, 10 ~ 15ml of enzyme solution is added, shakes 3 at 30~32 DEG C
~4h is filtered with sterile G2 funnel, collects filtrate in test tube, (5000 revs/min) removal enzyme solutions are centrifuged on centrifuge, heavy
It is 0.6mol/L MgSO that concentration is added in shallow lake4Sterile liquid is diluted to 1 × 103A/ml, as Protoplast suspension；
4. protoplast regeneration
4.1 regeneration culture medium production methods: regeneration culture medium formula is 180 ~ 200g of potato, 100 ~ 110g of sucrose, agar
20g, water 1000ml, peeling potatoes are cut into slices, and are put into pot, water 1000ml are added, heating is boiled 30 minutes, with 4 layers of yarn
Cloth filtering, is added agar in filtrate, continues heating and boils until agar melts completely, sucrose, heating stirring is added in fusion
Melt to sucrose, be eventually adding water and be settled to 1000ml, measures culture medium 200ml and be packed into the triangular flask of 250 ml, with sealing
Film sealing, sterilization treatment 30 minutes, culture medium is poured into sterile petri dish at 121 DEG C, in culture dish culture base unit weight be 20 ~
25ml is plating medium after cooling；
4.2 inoculated and cultured methods: taking 0.1ml Protoplast suspension to be put on plating medium, with glass bar by protoplast
Smearing is dispersed in plating medium surface, then, is put into culture in 20 ~ 25 DEG C of incubator, cultivates 8-10 days, in culture base table
It looks unfamiliar after growing white colony, takes single bacterium colony with inoculating tool, be put on slant medium (culture medium prescription is with 1.1),
It is cultivated at 20 ~ 25 DEG C, growing is Protoplast regeneration strains after mycelium；
5. monokaryon protoplast is identified
It takes mycelium to be placed on glass slide, drips water, covered is observed under the microscope, without clamp connection on mycelia
Label is monokaryon Protoplast fusion strain.
3. the method according to claim 1 using monokaryon Protoplast Technique breeding needle mushroom excellent variety, feature
Be: cultivation fruiting method includes 1. culture medium prescriptions and sterilizing methods；2. inoculated and cultured method；3. fruiting method；Specifically:
1. culture medium prescription and sterilizing methods
Cotton seed hulls or corncob 78%, wheat bran or rice bran 30%, calcium carbonate 2%, water content 65% prepare culture medium by formula rate,
It is then charged into 1100ml plastic bottle, sterilization treatment 2 hours at 121 DEG C；
2. inoculated and cultured method
Strain is accessed under gnotobasis in transfer room, bacterium germination is cultivated at 20-22 DEG C, is cultivated 23-25 days, mycelial growth is full
After bottle, into management of producing mushroom；
3. fruiting method
The strain that mycelial growth expires bottle is moved on in cultivating chamber, cap removing is gone, digs out surface layer strain, 10 ~ 15 DEG C of temperature, air
Relative humidity 95%, 10 ~ 50 lux of intensity of illumination reach 14cm ~ 16cm by 22 ~ 24 days acupuncture needle massee fruiting bodies length
Harvesting.
4. the method according to claim 1 using monokaryon Protoplast Technique breeding needle mushroom excellent variety, feature
It is: tissue isolated culture method specifically: take fructification to carry out tissue separation, cut stem and cap junction with scalpel
Bit organization is placed in PDA culture medium, is cultivated at 25 DEG C, after culture 5 days, selects the strain vigorous without miscellaneous bacteria, mycelial growth
It carries out switching culture, transfers on PDA slant medium, cultivate 8 ~ 10 days at 25 DEG C, as the strain that is separately cultured of tissue.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201810878454.7A CN109006189B (en) | 2018-08-03 | 2018-08-03 | Method for breeding needle mushroom fine variety by using mononuclear protoplast technology |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201810878454.7A CN109006189B (en) | 2018-08-03 | 2018-08-03 | Method for breeding needle mushroom fine variety by using mononuclear protoplast technology |
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| CN109006189A true CN109006189A (en) | 2018-12-18 |
| CN109006189B CN109006189B (en) | 2021-03-30 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109112070A (en) * | 2018-08-03 | 2019-01-01 | 四川省农业科学院土壤肥料研究所 | One plant of Strains of Flammulina velutipes |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1566340A (en) * | 2003-06-10 | 2005-01-19 | 四川省农业科学院土壤肥料研究所 | New method for edible mushroom distant protoplast fusion breeding |
| CN104956914A (en) * | 2015-06-18 | 2015-10-07 | 唐伟 | Breeding method for natural ganoderma lucidum |
| CN106244469A (en) * | 2016-09-28 | 2016-12-21 | 四川省农业科学院土壤肥料研究所 | A kind of method of Flammulina velutiper (Fr.) Sing unicellular selection-breeding strain excellent |
-
2018
- 2018-08-03 CN CN201810878454.7A patent/CN109006189B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1566340A (en) * | 2003-06-10 | 2005-01-19 | 四川省农业科学院土壤肥料研究所 | New method for edible mushroom distant protoplast fusion breeding |
| CN104956914A (en) * | 2015-06-18 | 2015-10-07 | 唐伟 | Breeding method for natural ganoderma lucidum |
| CN106244469A (en) * | 2016-09-28 | 2016-12-21 | 四川省农业科学院土壤肥料研究所 | A kind of method of Flammulina velutiper (Fr.) Sing unicellular selection-breeding strain excellent |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109112070A (en) * | 2018-08-03 | 2019-01-01 | 四川省农业科学院土壤肥料研究所 | One plant of Strains of Flammulina velutipes |
| CN109112070B (en) * | 2018-08-03 | 2021-10-08 | 四川省农业科学院土壤肥料研究所 | Flammulina velutipes strain |
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| CN109006189B (en) | 2021-03-30 |
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