CN112655463B - Process for deep culture of W192 agaricus bisporus liquid stock in fermentation tank - Google Patents
Process for deep culture of W192 agaricus bisporus liquid stock in fermentation tank Download PDFInfo
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Abstract
A preparation process for preparing W192 mushroom liquid stock seeds in batches through deep culture in a fermentation tank aims to prepare W192 liquid stock seeds with strong activity and high purity in batches, and the W192 liquid stock seeds can replace solid stock seeds to be used as seed sources for producing respiratory bag mushroom cultivation seeds. The invention relates to the technical field of edible fungus strains. The method comprises the following process flows: preparing a shake flask seed culture medium, sterilizing, cooling, inoculating, shake culturing, preparing a fermentation tank liquid stock culture medium, sterilizing, cooling, inoculating, deep culturing in a fermentation tank, inoculating a fungus bag (bottle), and culturing by fermentation. The W192 liquid stock prepared by the method has the advantages of high production efficiency, short culture period, low cost, strong activity, consistent bacterial age and the like.
Description
Technical Field
The invention belongs to the technical field of edible fungus strains, and particularly relates to a preparation process of an edible fungus liquid strain.
Background
The agaricus bisporus is edible fungus produced and consumed worldwide, the annual yield of the agaricus bisporus in China is more than 50% of the world, and the yield value is more than 100 hundred million yuan. In recent years, due to the increase of labor cost, the industrial cultivation of agaricus bisporus becomes a trend of mushroom cultivation and development, and the strain can be called as a 'chip' of the mushroom, which is a key for determining the production safety of the mushroom. The edible fungus institute (the inventor unit) of the agricultural academy of Fujian province is the only scientific research unit in China for focusing on the research and development of the agaricus bisporus strain breeding and seed production process, and the bred As2796 mushroom strain is the main cultivated variety (more than 90 percent) for domestic mushroom cultivation for twenty years, and is suitable for the traditional artificial cultivation mode. In order to meet the requirement of rapid development of the current industrial mushroom planting, the inventor breeds a new W192 mushroom strain in 2012 of the institute, has the characteristics of high yield, high quality, obvious tide and the like, is relatively suitable for domestic industrial planting, can compete with overseas industrial varieties and has certain advantages. Through the development of new technology (patent number: ZL 200910111583.4) for preparing respiratory bag mushroom cultivars by adopting solid stock seeds and the preparation technology (patent number: ZL 201310365188.5) for shaking bottle culture of W2000 liquid stock seeds, along with the popularization of W192 strains in domestic mushroom industrial production, the development of technology for preparing W192 liquid stock seeds in batch by adopting deep culture of a fermentation tank is needed to meet the requirement of W192 strain batch production. Therefore, in 2017, the research of application of agaricus bisporus liquid stock in producing culture seeds of breathable bags (special for scientific research institutes of public welfare) is carried out by China, and the project group researches for 3 years, a new process for deep culturing W192 liquid stock by adopting a fermentation tank is developed, and an invention patent is applied to form protection of a technical closed loop.
Disclosure of Invention
The invention aims to provide a process for preparing W192 agaricus bisporus liquid stock seeds in batches by deep culture in a fermentation tank, and the process can prepare W192 liquid stock seeds with strong vitality in batches for producing respiratory bag cultivators, so that the requirements of industrial planting enterprises on W192 strains are met. The W192 liquid stock seed prepared by the process has the characteristics of high yield and strong vitality, and can further improve the production efficiency and the process level of the respiratory bag cultivar and reduce the production cost when being used for producing the respiratory bag cultivar.
The invention is specifically described as follows:
1 Process flow
Preparing a shake flask seed culture medium, sterilizing, cooling, inoculating, shake culturing, preparing a fermentation tank liquid stock culture medium, sterilizing, cooling, inoculating, deep culturing in a fermentation tank, inoculating a fungus bag (bottle), and culturing by fermentation.
2 Medium formulation
2.1 W192 mother liquor culture medium
Millet flour 7.5g/L, soybean flour 5.0g/L, glucose 10g/L, peptone 2g/L and MgSO 4 ·7H 2 O 0.75g/L, KH 2 PO 4 2.0g/L。
2.2W192 fermentation tank liquid stock culture medium
2.2.1 millet flour 30g/L, soybean flour 12g/L, peptone 3g/L, KH 2 PO 4 1.25g/L，MgSO 4 ·7H 2 O 0.75g/L，pH 6.0。
2.2.2 corn flour 30g/L, soybean flour 12g/L, peptone 3g/L, KH 2 PO 4 1.25g/L，MgSO 4 ·7H 2 O0.75 g/L, pH 6.0.2.3 cultivar Medium
2.3.1 wheat 94%, gypsum powder 3%, light calcium carbonate 3%, water content 48% + -1%, pH 7.5-8.0.
94% of millet, 3% of gypsum powder, 3% of light calcium carbonate, 48% +/-1% of water content and 7.5-8.0 of pH.
3. Preparation of the culture Medium
3.1 preparation of shake flask seed Medium
Accurately weighing millet flour 7.5g and soybean flour 5.0g, placing into a stainless steel pot, adding 500mL of water, boiling for 5min, adding glucose 10g, peptone 2g and MgSO 4 ·7H 2 O 0.75g，KH 2 PO 4 2.0 And g, uniformly stirring the mixture by using a glass rod, fixing the volume to 1000mL by using a cylinder to obtain a shake flask seed culture medium, placing the prepared culture medium on a magnetic stirrer for split charging, packaging 100mL of each triangular flask, placing the triangular flasks in a sterilizer at 121 ℃, sterilizing for 30min, and naturally cooling the sterilized culture medium for later use.
3.2 preparation of liquid stock Medium in fermenter
According to the formula, a corresponding amount of fermentation tank liquid stock culture medium is weighed, placed in a stainless steel pot, 3L of water is added, and the mixture is stirred uniformly, thus obtaining a culture medium concentrated solution. When the water temperature of 97L in the fermentation tank reaches 80 ℃, an air pump is started for ventilation, the prepared culture medium concentrated solution is poured into the fermentation tank, 20mL of dichlord is added, the fermentation tank is sealed, sterilization is carried out for 30min at 121 ℃, and cooling is carried out, thus obtaining the fermentation tank liquid stock culture medium.
3.3 liquid stock culture Medium Sterilization operation of fermenter
3.3.1 adding Water to the tank Sandwich
Firstly, adding water into an interlayer of a liquid fermentation tank, and the method comprises the following steps: the silica gel pipe is used for connecting the water pipe and the interlayer water inlet valve, the interlayer water outlet valve is opened for adding water (so as to prevent the interlayer pressure from being excessively high and damaging the inner tank body), and the water is added to the interlayer water outlet valve for discharging water.
3.3.2 liquid Medium Sterilization
The temperature of the temperature control box is adjusted to 121 ℃, the heating rod is opened to start heating, the interlayer water drain valve is opened, and the virtual pressure and the redundant water in the interlayer are discharged.
3.3.2.1 adding water to the tank
Water is added into the tank body through an inoculation valve or an inoculation port, and is added below the tank body sight glass until the water temperature rises to 60 ℃.
3.3.2.2 adding into culture medium
When the water temperature in the tank body reaches more than 60 ℃, adding the prepared liquid stock culture medium into the tank body, preferably adding 20mL of defoaming agent when the liquid height is higher than a tank body observation mirror, then screwing the tank cover of the fermentation tank, opening an air pump, and opening the air pump: opening a filter shell air release valve, opening a filter shell air inlet valve, opening an air pump, closing the filter shell air release valve, opening a tank air release valve until the temperature rises to 70 ℃, and closing the air pump, wherein the steps are as follows: guan Qibeng, closing the filter shell air release valve and the air inlet valve, and closing the tank air release valve. And heating is continued.
3.3.3 liquid Medium Sterilization
When the interlayer water discharge valve discharges hot steam for 3-5min, the interlayer air discharge valve is closed (the two ends of the steam pipe are connected with the interlayer air discharge valve and the tank air inlet valve, the one-way valve arrow is downward), when the interlayer pressure rises to 0.1MPa, the interlayer air discharge valve is opened, the tank air inlet valve is opened again, the tank air discharge valve is opened, and after the air inlet pipe burns hands, the tank air discharge valve is closed. When the pressure of the tank body is expressed to 0.15MPa, starting timing, opening a tank body air release valve for 1/3, regulating the temperature according to the pressure, and maintaining for 30min for sterilization.
3.3.4 Cooling
After timing, closing the tank air inlet valve and the Guan Gaceng air outlet valve, regulating the temperature of the temperature control meter to 25 ℃, and closing the heating rod.
3.3.5 placing Sandwich Hot Water
At this time, the air release valve of the tank body can be opened again to a large point, and the interlayer hot water is discharged through the interlayer water inlet valve until the pressure of the interlayer pressure gauge is reduced to 0.
3.3.6 liquid Medium Cooling
And opening an interlayer water drain valve to connect the water drain pipe to a sewer, and connecting the interlayer water inlet valve to the water pipe through a silicone pipe for cooling. When the pressure of the tank pressure gauge is reduced to 0.05MPa, the air pump is turned on (preventing pollution caused by negative pressure generated in the cooling process of the tank and enabling cold water at the lower part to be cooled upwards more quickly). And (3) opening the air pump: and (3) opening a filter shell air release valve, opening a filter shell air inlet valve, opening an air pump, closing the filter shell air release valve (condensate water can be discharged through a small filter shell air release valve in the culture process), regulating the pressure of a tank body to be more than 0MPa and less than 0.05MPa through a tank body air release valve, and reducing the temperature to 24 ℃ to start inoculation.
3.4 preparation of cultivar Medium
Wheat and millet matrix culture medium is prepared by a V-shaped digester according to a cultivar formula; and (3) accurately weighing the granular substrate culture medium according to a formula, uniformly stirring in a stirrer, bagging in a container which is a breathable bag, sterilizing at 126 ℃ for 2.5h, and naturally cooling to obtain the solid culture medium for the cultivar.
4 activation of the seed
From the deposited slant agaricus bisporus strain, under aseptic operation condition, 0.2cm×0.5cm size bacterial block is cut out, inoculated on PDA plate, placed in constant temperature incubator, culture temperature is 24 deg.C, and cultured for 20d.
5 liquid spawn culture
5.1 shake flask seed culture
Cutting the activated agaricus bisporus plate mother strain into bacterial blocks with the diameter of 1.0 and cm by using a puncher, transferring the bacterial blocks to 100mL sterilized shake flask seed culture medium in 250mL triangular flasks, and inoculating 2 bacterial blocks into each triangular flask; then culturing for 8d by rotary shaking on a culture shaker with the culture temperature of 24 ℃ and the shaking speed of 180r/min to obtain agaricus bisporus shaking flask seeds.
5.2 submerged culture of liquid stock in fermenter
Winding gauze with iron wire ring with handle, pouring 95% alcohol. The flame ring is completely sleeved on the inoculation port, the air release valve is opened, the flame ring is ignited, the inoculation port is opened, the tank air release valve is closed when the inoculation port is opened to half, the inoculation port is completely opened, then strains are quickly, stably and lightly connected (the tank pressure can rise again in the process of opening the inoculation port, and the tank air can be released again through the tank air release valve), and the inoculation port is screwed down. Opening the air release valve of the tank body, adjusting the pressure to be more than 0 and less than 0.05MPa, and entering the culture stage. Deep fermentation culture is carried out for 7d to obtain agaricus bisporus liquid stock.
6 observing
After 24h inoculation, the strain germination and growth were observed every 12 th h. Five general looks and smell: looking at the dissolved oxygen meter, the oxygen content of the normal culture liquid of the fermentation tank is maintained at 7.5-9.5mg/L, and if the oxygen content of the fermentation liquid is lower, the fermentation liquid is indicated to be polluted; looking at a pH meter instrument, the pH of the fermentation liquor is maintained between 4.8 and 6.5 in the whole process of submerged culture in the fermentation tank; and thirdly, the color of the bacterial liquid is seen, and the color of the normal bacterial liquid is pure. Although the color of light yellow, light brown and the like is present, the color is not turbid; fourthly, the clarity of the bacterial liquid is seen, the early stage of the culture is slightly turbid, and no fine particles and floccules exist in the bacterial liquid at the later stage of the culture, so that the bacterial liquid is more and more clear and transparent, otherwise, the bacterial liquid is abnormal; fifthly, whether burrs around the fungus balls are obvious or not and whether the number of the fungus balls is increased or not is judged. The concentration of the bacteria balls increases rapidly in the range of 48-72 and h, and the color depth indicates that the bacteria balls are bad if the bacteria balls become turbid, milder and have wine taste. The smell is that the smell of the fungus liquid is smelled, the sweet smell of the culture liquid becomes lighter along with the long culture time, and only a light fungus liquid has fresh fragrance in the later period.
7 fungus-collecting bag (bottle) and culture
The agaricus bisporus stock with corresponding weight is inoculated into a fungus bag (bottle) by using the pressure of a fermentation tank in an ultra-clean workbench through an inoculation gun. After inoculation, the mycelia are placed in a culture room for culture at 24 ℃, and the germination and bacteria removal conditions of the mycelia are observed every 24 hours.
The invention has the advantages that:
1) Compared with shaking flask culture, the method has high production efficiency and can be used for mass production.
2) And solid stock is shorter than the culture time (40-50 days is shortened to 6-7 days)
3) Strong activity and consistent bacterial age.
4) Avoid the problems of hidden pollution and the like, and has high yield (more than 98 percent).
5) Simple inoculation, easy automation operation and low cost.
Drawings
FIG. 1 is a flow chart of the preparation process of the present invention
The specific embodiment is as follows:
example 1:
1. preparation of W192 mother liquor Strain 1L
a. Accurately weighing 7.5g of millet powder, 5g of soybean powder, 2g of peptone, 10g of glucose and KH respectively 2 PO 4 2g ，MgSO 4 ·7H 2 O 0.75g ；
b. Adding 0.1L distilled water into millet powder and peptone, mixing, and diluting to 0.5L.
c. Adding 0.5L distilled water into soybean powder, heating to boil, filtering with four layers of gauze after 20min, and discarding yellow residue to obtain soybean powder filtrate for use.
d. Mixing the liquids obtained in step b and c, and adding 2g KH 2 PO 4 And 0.75g MgSO 4 ·7H 2 O, uniformly stirring, pouring into a 1L measuring cylinder, and supplementing the mixed solution to 1L by using distilled water to obtain a mother solution culture medium;
e. 100mL of mother liquor culture medium with the volume of 100mL of the measuring cylinder is poured into a 250mL triangular flask, and a silica gel plug is plugged at the bottle mouth;
f. sterilizing with 0.1mpa and 121 deg.c high pressure steam for 20min;
g. cooling the mother liquor culture medium, and then placing the mother liquor culture medium in an ultra-clean workbench for sterilization by irradiation of an ultraviolet lamp for 30min;
h. under the aseptic condition, inoculating the agaricus bisporus W192 fungus blocks with the size of 0.5cm multiplied by 0.5cm on the PDA inclined plane into a mother liquor culture medium, and fermenting and culturing at the culture temperature of 25 ℃ by a 180r/min rotary oscillator for 8d to finish the preparation of W192 mother liquor strains.
2. Preparation of W192 fermentation tank liquid stock culture medium
After the W192 mother liquor strain is prepared, the preparation of the W192 fermentation tank liquid stock culture medium is carried out, and the method comprises the following steps:
a. weighing 30g of millet powder, 12g of soybean powder, 3g of yeast powder and KH per liter of liquid stock culture medium 2 PO 4 1.25g，MgSO 4 ·7H 2 O,0.75g。
b. Accurately weighing millet powder, soybean powder, yeast powder and KH according to 40L liquid stock formula 2 PO 4 And MgSO 4 ·7H 2 O, adding 4L of water, uniformly mixing to obtain a culture medium concentrated solution, pouring the culture medium concentrated solution into a fermentation tank with the capacity of 100L, adding 20mL of enemy, adding water for dilution until the water temperature in the fermentation tank reaches 80 ℃, starting an air pump for ventilation, pouring the prepared culture medium concentrated solution into the fermentation tank, adding enemy, and sealing the fermentation tank.
c. Sterilizing the culture solution at 121 deg.C under 0.1mpa for 20min according to the operation procedure of fermentation tank, and cooling to 24 deg.C;
d. under the protection of flame, 400mL of liquid seeds are inoculated into the liquid stock culture medium of the fermentation tank;
e. aeration rate after inoculation is 1.5m 3 And/min, fermenting and culturing for 7d to finish the preparation of the liquid stock of the W192 fermentation tank.
3. W192 breathing bag cultivar production
Sterile operation is carried out in hundred-grade clean space, the liquid stock of the W192 fermentation tank is inoculated into a culture medium of a respiratory bag cultivation seed, and the culture is carried out for 15 days at 25 ℃, so that the yield can reach 99%.
Example 2:
1. preparation of W192 mother liquor Strain 1L
a. 25g corn flour, 10g soybean powder, peptone 2g, glucose 10g and KH were accurately weighed respectively 2 PO 4 1.75g ，MgSO 4 ·7H 2 O 0.75g ；
b. Corn flour and peptone are added with 0.1L distilled water to be uniformly mixed and then diluted to 0.5L mixed solution.
c. Adding 0.5L distilled water into soybean powder, heating to boil, filtering with four layers of gauze after 20min, and discarding yellow residue to obtain soybean powder filtrate for use.
d. Mixing the liquids obtained in step b and c and adding 1.75g KH 2 PO 4 And 0.75g MgSO 4 ·7H 2 O, uniformly stirring, pouring into a 1L measuring cylinder, and supplementing the mixed solution to 1L by using distilled water to obtain a mother solution culture medium;
e. 100mL of mother liquor culture medium with the volume of 100mL of the measuring cylinder is poured into a 250mL triangular flask, and a silica gel plug is plugged at the bottle mouth;
f. sterilizing with 0.1mpa and 121 deg.c high pressure steam for 20min;
g. cooling the mother liquor culture medium, and then placing the mother liquor culture medium in an ultra-clean workbench for sterilization by irradiation of an ultraviolet lamp for 30min;
h. under the aseptic condition, inoculating the W192 fungus blocks of the agaricus bisporus with the size of 0.5cm multiplied by 0.5cm on the PDA inclined plane into a mother liquor culture medium, and fermenting and culturing for 7d at the culture temperature of 25 ℃ by a 160r/min rotary oscillator to finish the preparation of the W192 mother liquor strain.
2. Preparation of W192 fermentation tank liquid stock culture medium
After the W192 mother liquor strain is prepared, the preparation of the W192 fermentation tank liquid stock culture medium is carried out, and the method comprises the following steps:
a. 30g of corn meal, 12g of soybean meal, 3g of peptone and KH are weighed according to each liter of liquid stock culture medium 2 PO 4 1.25g，MgSO 4 ·7H 2 O,0.75g。
b. Precisely weighing corn flour, soybean powder, peptone and KH according to 40L liquid stock formula 2 PO 4 And MgSO 4 ·7H 2 O, adding 4L of water, mixing uniformly to obtain culture medium concentrated solution, pouring into a fermentation tank with the capacity of 100L, and adding water to dilute to 40L of culture solution. Adding 20mL of dichlord, adding water to dilute to 40L of culture solution, starting an air pump to ventilate when the water temperature in the fermentation tank reaches 80 ℃, pouring the prepared culture medium concentrated solution into the fermentation tank, adding dichlord, and sealing the fermentation tank.
c. Sterilizing the culture solution at 121 deg.C under 0.1mpa for 20min according to the operation procedure of fermentation tank, and cooling to 24 deg.C;
d. under the protection of flame, 400mL of liquid seeds are inoculated into the liquid stock culture medium of the fermentation tank;
e. aeration rate after inoculation is 1.5m 3 And/min, fermenting and culturing for 7d to finish the preparation of the liquid stock of the W192 fermentation tank.
3. W192 breathing bag cultivar production
Sterile operation is carried out in hundred-grade clean space, the liquid stock of the W192 fermentation tank is inoculated into a culture medium of a respiratory bag cultivation seed, and the culture is carried out for 15 days at 25 ℃, so that the yield can reach 99%.
Claims (2)
1. A method for deep culture of W192 agaricus bisporus liquid stock in a fermentation tank is characterized by comprising the following steps:
1) Preparation of W192 mother liquor strain
Cutting the activated agaricus bisporus plate mother strain into bacterial blocks with the diameter of 1.0 and cm by using a puncher, transferring the bacterial blocks to 100mL of sterilized W192 mother solution culture medium in 250mL triangular flasks, and inoculating 2 bacterial blocks to each triangular flask; then, culturing for 8d by rotary shaking on a culture shaker with the culture temperature of 24 ℃ and the shaking flask revolution of 180r/min to obtain agaricus bisporus mother liquor strain;
2) Submerged culture of liquid stock in fermentation tank
Winding gauze with iron wire ring with handle, pouring 95% alcohol; the flame ring is completely sleeved on the inoculation port, the air release valve is opened, the flame ring is ignited, the inoculation port is opened, the tank air release valve is closed when the inoculation port is opened to half, the agaricus bisporus mother liquor strain is quickly, stably and lightly connected into the liquid stock culture medium of the sterilization fermentation tank after the inoculation port is completely opened, and the inoculation port is screwed; opening a tank body air release valve, adjusting the pressure to be more than 0 and less than 0.05MPa, and entering a culture stage; deep fermentation culture is carried out for 7d to obtain agaricus bisporus liquid stock;
the preparation method of the liquid stock culture medium of the fermentation tank comprises the following steps:
(1) Weighing 30g of corn powder or millet powder, 12g of soybean powder, 3g of yeast powder and KH per liter of liquid stock culture medium 2 PO 4 1.25g，MgSO 4 ·7H 2 0.75g of O is prepared;
(2) Weighing raw materials of a liquid stock culture medium of a fermentation tank according to a formula, placing the raw materials in a stainless steel pot, adding 4L of water, and uniformly stirring to obtain a culture medium concentrated solution; diluting with water to obtain culture solution, and sterilizing to obtain fermentation tank liquid stock culture medium;
the sterilization method of the liquid stock culture medium of the fermentation tank comprises the following steps:
1) Tank interlayer water adding
Firstly, adding water into an interlayer of a liquid fermentation tank, and the method comprises the following steps: the silica gel pipe is used for connecting the water pipe and the interlayer water inlet valve, the interlayer water outlet valve is opened to add water so as to prevent the interlayer pressure from being excessively high and damaging the inner tank body, and the water quantity is added to the water outlet of the interlayer water outlet valve;
2) Sterilization of liquid culture medium
Adjusting the temperature of the temperature control box to 121 ℃, opening a heating rod to start heating, opening an interlayer water drain valve, and discharging virtual pressure and redundant water in the interlayer;
2.1 Water is added into the tank body
Adding water into the tank body through an inoculation valve or an inoculation port, and adding water to the position below the tank body sight glass until the water temperature rises to 60 ℃;
2.2 Adding a culture medium
When the water temperature in the tank body reaches more than 60 ℃, adding the prepared culture solution into the tank body, adding 20mL of defoaming agent into the tank body, screwing the tank cover of the fermentation tank, opening an air pump, and opening the air pump: opening a filter shell air release valve, opening a filter shell air inlet valve, opening an air pump, closing the filter shell air release valve, opening a tank air release valve until the temperature rises to 70 ℃, and closing the air pump, wherein the steps are as follows: guan Qibeng, closing the filter shell air release valve and the air inlet valve, and closing the tank air release valve; continuously heating;
2.3 Sterilization of liquid culture medium
When the interlayer water discharge valve discharges hot steam for 3-5min, closing the interlayer air outlet valve, when the pressure of the interlayer rises to 0.1MPa, opening the interlayer air outlet valve, then opening the tank air inlet valve, opening the tank air outlet valve slightly, and closing the tank air outlet valve after the air inlet pipe burns hands; when the pressure of the tank body is expressed to 0.15MPa, starting timing, opening a tank body air release valve for 1/3, regulating the temperature according to the pressure, and maintaining for 30min for sterilization;
2.4 Temperature reduction)
After timing, closing the tank air inlet valve and the Guan Gaceng air outlet valve, regulating the temperature of the temperature control meter to 25 ℃, and closing the heating rod;
2.5 Hot water in interlayer
Opening a large air release valve, and discharging interlayer hot water through an interlayer water inlet valve until the pressure of an interlayer pressure gauge is reduced to 0;
2.6 Liquid culture medium cooling
Opening an interlayer water drain valve to connect the water drain pipe to a sewer, and connecting the interlayer water inlet valve to the water pipe through a silicone pipe for cooling; when the pressure of the tank pressure gauge is reduced to 0.05MPa, the air pump is opened, so that pollution caused by negative pressure generated in the cooling process of the tank is prevented, the lower cold water is cooled upwards faster, and the air pump is opened: and (3) opening a filter shell air release valve, opening a filter shell air inlet valve, opening an air pump, closing the filter shell air release valve, discharging condensed water through the small filter shell air release valve in the culture process, regulating the pressure of a tank body to be 0-0.05 MPa through the tank body air release valve, and reducing the temperature to 24 ℃ to start inoculation.
2. The culture method according to claim 1, wherein the preparation method of the W192 mother liquor medium comprises the steps of:
accurately weighing millet flour 7.5g and soybean flour 5.0g, placing into a stainless steel pot, adding 500mL of water, boiling for 5min, adding glucose 10g, peptone 2g and MgSO 4 ·7H 2 O 0.75g，KH 2 PO 4 2.0g of the culture medium is stirred uniformly by a glass rod, the volume of the culture medium is fixed to 1000mL by a cylinder to obtain a shake flask seed culture medium, the prepared culture medium is placed on a magnetic stirrer for split charging, 100mL of each triangular flask is placed in a sterilizer, the sterilization is carried out for 30min at 121 ℃, and the sterilized culture medium is naturally cooled for standby.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1651568A (en) * | 2004-02-03 | 2005-08-10 | 李勇 | Edible fungus liquid culture submerged fermentation technology |
| CN103782801A (en) * | 2013-12-26 | 2014-05-14 | 朝阳鑫源农副产品开发有限公司 | Agaricus Bisporus liquid spawn and preparation method thereof |
| CN103875446A (en) * | 2013-10-24 | 2014-06-25 | 江西省农业科学院农业应用微生物研究所 | Method of producing Agrocybe aegerita strains by liquid culture and preservation method of Agrocybe aegerita strain |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100789273B1 (en) * | 2006-05-25 | 2008-01-02 | 씨제이 주식회사 | A method for culturing Agaricus bisporus mycellium and a medium for culturing the same |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1651568A (en) * | 2004-02-03 | 2005-08-10 | 李勇 | Edible fungus liquid culture submerged fermentation technology |
| CN103875446A (en) * | 2013-10-24 | 2014-06-25 | 江西省农业科学院农业应用微生物研究所 | Method of producing Agrocybe aegerita strains by liquid culture and preservation method of Agrocybe aegerita strain |
| CN103782801A (en) * | 2013-12-26 | 2014-05-14 | 朝阳鑫源农副产品开发有限公司 | Agaricus Bisporus liquid spawn and preparation method thereof |
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