CN112655463A - Process for submerged culture of W192 agaricus bisporus liquid stock seeds in fermentation tank - Google Patents
Process for submerged culture of W192 agaricus bisporus liquid stock seeds in fermentation tank Download PDFInfo
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Abstract
A preparation process for preparing W192 mushroom liquid stock in batches by submerged culture of a fermentation tank aims at preparing W192 liquid stock with strong activity and high purity in batches, and can replace solid stock to be used as a seed source for producing breathing bag mushroom cultivated species. The invention relates to the technical field of edible fungus strains. Comprises the following process flows: the preparation of the seed culture medium by shaking the flask → sterilization → cooling → inoculation → shake cultivation → the preparation of the liquid stock culture medium of the fermentation tank → sterilization → cooling → inoculation → submerged cultivation of the fermentation tank → inoculation bag (bottle) → spawn cultivation. The method for preparing the W192 liquid breeder seeds has the advantages of high production efficiency, short culture period, low cost, strong activity, consistent fungus 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 which is produced and consumed in the world, the annual output of the agaricus bisporus in China accounts for more than 50 percent of the world, and the output value is more than 100 billion yuan. In recent years, due to the increase of labor cost, the industrial planting of agaricus bisporus becomes the trend of mushroom planting development, and the strain can be called as a mushroom chip, which is the key for determining the production safety of mushrooms. The edible fungus research institute (inventor unit) of agricultural academy of sciences of Fujian province is the only domestic research and development unit dedicated to the breeding and seed production process of agaricus bisporus strains, and the bred As2796 mushroom strain is the main cultivated variety (accounting for more than 90 percent) of 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 current mushroom industrial planting, a new W192 mushroom strain is bred in 2012, has the characteristics of high yield, high quality, obvious tide and the like, is suitable for domestic industrial planting, can compete with foreign industrial varieties and has certain advantages. After years of research and development of a new process (patent number: ZL 200910111583.4) for preparing breathing bag mushroom cultivars by adopting solid breeder seeds and a preparation process (patent number: ZL 201310365188.5) for shaking culture of W2000 liquid breeder seeds, along with popularization of W192 strains in domestic mushroom industrial production, a seed production process for preparing W192 liquid breeder seeds in batches by adopting submerged culture of a fermentation tank is urgently needed to meet the requirement of batch production of the W192 strains. Therefore, the 2017 Fujian province science and technology hall establishes an application research of the agaricus bisporus liquid stock in the production of the breathable bag cultivated species (special item of public welfare scientific research institutions) and is born by the inventor, a new process for deeply culturing the W192 liquid stock by adopting a fermentation tank is developed by a project group through 3 years of research, and the invention patent is applied to form the protection of the technical closed loop.
Disclosure of Invention
The invention aims to provide a process for preparing W192 agaricus bisporus liquid stock seeds in batches by adopting submerged culture of a fermentation tank, and the process can prepare the W192 liquid stock seeds with strong activity in batches for producing respiration bag cultivated species, and meets the requirements of factory planting enterprises on the W192 strains. The W192 liquid stock prepared by the process has the characteristics of high yield and strong activity, and can further improve the production efficiency and the process level of the breathing bag cultivated species and reduce the production cost when used for the production of the breathing bag cultivated species.
The invention is specifically described as follows:
1 Process flow
The preparation of the seed culture medium by shaking the flask → sterilization → cooling → inoculation → shake cultivation → the preparation of the liquid stock culture medium of the fermentation tank → sterilization → cooling → inoculation → submerged cultivation of the fermentation tank → inoculation bag (bottle) → spawn cultivation.
2 culture medium formula
2.1W 192 mother liquor culture medium
7.5g/L millet powder, 5.0g/L soybean powder, 10g/L glucose, 2g/L peptone and MgSO 24·7H2O 0.75g/L, KH2PO4 2.0g/L。
2.2W192 fermenter liquid stock culture medium
2.2.1 g/L millet powder, 12g/L soybean powder, 3g/L peptone and KH2PO4 1.25g/L，MgSO4·7H2O 0.75g/L，pH 6.0。
2.2.2 g/L corn flour 30g/L soybean flour 12g/L peptone 3g/L KH2PO4 1.25g/L，MgSO4·7H2O0.75 g/L, pH 6.0. 2.3 cultivar Medium
94 percent of wheat 2.3.1, 3 percent of gypsum powder, 3 percent of light calcium carbonate, 48 percent +/-1 percent of water content and 7.5 to 8.0 of pH value.
94% of millet 2.3.2, 3% of gypsum powder, 3% of light calcium carbonate, 48% +/-1% of water content and 7.5-8.0 of pH value.
3 preparation of the Medium
3.1 preparation of Shake flask seed Medium
Accurately weighing 7.5g of millet flour and 5.0g of soybean flour, placing in a stainless steel pot, adding 500mL of water, boiling for 5min, adding 10g of glucose, 2g of peptone and MgSO 24·7H2O 0.75g，KH2PO42.0 g of the seed culture medium is stirred uniformly by a glass rod, the volume of the seed culture medium is fixed to 1000mL by a measuring cylinder to obtain a shake flask seed culture medium, and the prepared culture medium is placedSubpackaging in magnetic stirrer, bottling in 100mL bottles, sterilizing in sterilizer at 121 deg.C for 30min, and naturally cooling the sterilized culture medium.
3.2 preparation of liquid stock culture Medium for fermenter
According to the formula, weighing a corresponding amount of fermentation tank liquid stock culture medium, placing the fermentation tank liquid stock culture medium in a stainless steel pot, adding 3L of water, and uniformly stirring to obtain the culture medium concentrated solution. And (3) when the water temperature of 97L in the fermentation tank reaches 80 ℃, starting an air pump to ventilate, pouring the prepared culture medium concentrated solution into the fermentation tank, adding 20mL of foam, sealing the fermentation tank, sterilizing at 121 ℃ for 30min, and cooling to obtain the fermentation tank liquid stock culture medium.
3.3 Sterilization of the liquid stock culture Medium of the fermenter
3.3.1 tank body interlayer Water addition
Firstly, adding water into an interlayer of a liquid fermentation tank, wherein the method comprises the following steps: the silicone tube is connected with 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 overhigh to damage the inner tank body), and the water amount is added to the interlayer water outlet valve to obtain the water-saving interlayer water tank.
3.3.2 Sterilization of liquid Medium
The temperature of the temperature control box is adjusted to 121 ℃, the heating rod is opened to start heating, the water drain valve of the interlayer is opened, and the virtual pressure and the excess water in the interlayer are drained.
3.3.2.1 Water can
Adding water into the tank body through the inoculation valve or the inoculation port, and adding the water below a tank body sight glass until the water temperature rises to 60 ℃.
3.3.2.2 addition of 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, adding 20mL of defoaming agent when the liquid height is more than the observation mirror of the tank body, screwing down the fermentation tank cover, opening the air pump, and opening the air pump: opening a filter shell air release valve, opening a filter shell air inlet valve, then 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: the air pump is closed, the air release valve and the air inlet valve of the filter shell are closed, and then the air release valve of the tank body is closed. And continuously heating.
3.3.3 Sterilization of liquid Medium
When the hot steam is discharged from the interlayer water discharge valve for 3-5 min, 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 body air inlet valve, the arrow of the one-way valve is downward), when the interlayer pressure is increased to 0.1MPa, the interlayer air discharge valve is opened, the tank body air inlet valve is opened, the tank body air discharge valve is opened slightly, and the tank body air discharge valve is closed after the air inlet pipe is. And when the pressure gauge of the tank body reaches 0.15MPa, timing, opening a tank body air release valve 1/3, and adjusting the temperature according to the pressure to keep sterilization for 30 min.
3.3.4 Cooling
After the timing is finished, the air inlet valve of the tank body is closed, the air outlet valve of the interlayer is closed, the temperature of the temperature control meter is adjusted to 25 ℃, and the heating rod is closed.
3.3.5 interlayer hot water
At the moment, the tank body air release valve can be opened to a large point again, and interlayer hot water is released 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 the interlayer water drain valve to connect the water drain pipe to the sewer, and connecting the interlayer water inlet valve to the water inlet pipe through the silica gel pipe for cooling. When the pressure of the pressure gauge of the tank body is reduced to 0.05MPa, the air pump is started (the tank body is prevented from generating negative pressure in the cooling process to cause pollution, and the lower cold water is cooled upwards quickly). Starting an air pump: opening the air release valve of the filter shell, opening the air inlet valve of the filter shell, then opening the air pump, closing the air release valve of the filter shell (the condensed water can be discharged through the small air release valve of the filter shell in the culture process), adjusting the pressure of the tank body to be more than 0MPa and less than 0.05MPa through the air release valve of the tank body, and starting inoculation when the temperature is reduced to 24 ℃.
3.4 preparation of the culture Medium of the cultivars
Preparing a wheat and millet substrate culture medium by using a V-shaped digester according to a cultivar formula; and (3) accurately weighing the granular substrate culture medium according to the formula, putting the granular substrate culture medium into a stirrer, uniformly stirring, bagging the granular substrate culture medium in a ventilating bag, sterilizing at 126 ℃ for 2.5h, and naturally cooling to obtain the solid culture medium of the cultivated species.
4 bacterial activation
From the preserved strain of Agaricus bisporus, 0.2cm × 0.5cm of the strain block is cut out under aseptic conditions and inoculated on a PDA plate, and the plate is placed in a constant temperature incubator at 24 ℃ for 20 days.
5 liquid spawn culture
5.1 Shake flask seed culture
Cutting the activated agaricus bisporus plate mother seeds into bacterium blocks with the diameter of 1.0 cm by using a hole puncher, transferring the bacterium blocks to 100mL of sterile shake flask seed culture medium in a 250mL triangular flask, and inoculating 2 bacterium blocks to each triangular flask; then rotationally oscillating and culturing for 8d on a culture oscillator with the culture temperature of 24 ℃ and the oscillation rotation number of 180r/min to obtain the shake flask seeds of the agaricus bisporus.
5.2 submerged cultivation of liquid stock in fermenter
The gauze was wrapped with a wire loop with a handle and poured with 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 air release valve of the tank body is closed when the inoculation port is opened to a half, the strains are quickly, stably and lightly inoculated after the inoculation port is completely opened (the pressure of the tank body may rise again in the process of opening the inoculation port, and the air can be released again through the air release valve of the tank body), 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. And carrying out submerged fermentation culture for 7d to obtain an agaricus bisporus liquid stock.
6 Observation
After 24h of inoculation, the germination and growth of the strains are observed every 12 h. Generally, the five people see and smell one: at one glance, the oxygen content of the normal culture solution of the fermentation tank is maintained at 7.5-9.5mg/L by using a dissolved oxygen meter, and if the oxygen content of the fermentation solution is low, the fermentation solution is polluted; secondly, a pH meter is used, and the whole process of the fermentation broth for submerged culture in the fermentation tank is maintained between 4.8 and 6.5; the color of the bacteria liquid is seen, and the color of the normal bacteria liquid is pure. Although the color is light yellow, light brown, and the like, the color is not turbid; fourthly, the bacterial liquid is clear, slightly turbid in the early stage of culture and free of fine particles and floccules in the bacterial liquid in the later stage of culture, so that the bacterial liquid is more and more clear and transparent, otherwise, the bacterial liquid is abnormal; and fifthly, judging whether burrs around the bacteria balls are obvious and the number of the bacteria balls is increased. The concentration of the bacteria balls increases faster within 48-72 hours, and if the bacteria balls become turbid, musty and vinous, the color is dark, which indicates that the bacteria balls are bad. The smell of the bacteria liquid is smelled, the sweet taste of the culture solution becomes lighter along with the growth of the culture time, and only a light bacteria liquid faint scent is generated in the later period.
7 inoculating bag (bottle) and culturing
Inoculating agaricus bisporus stock seeds with corresponding weight into a fungus bag (bottle) through an inoculation gun by using the tank pressure of a fermentation tank in a clean bench. After inoculation, the mixture is placed in a culture room for culture at 24 ℃, and hypha germination and bacterium walking conditions are observed every 24 hours.
The invention has the advantages that:
1) and the production efficiency is higher than that of the shake flask culture, and the large-scale production can be realized.
2) Shorter than the solid stock culture (shortened to 6-7 days in 40-50 days)
3) Strong activity and consistent fungus age.
4) The problems of recessive pollution and the like are avoided, and the yield is high (more than 98%).
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 implementation mode is as follows:
example 1:
1. preparation of W192 mother liquor Strain 1L
a. Accurately weighing 7.5g millet powder, 5g soybean powder, 2g peptone, 10g glucose and KH2PO4 2g ，MgSO4·7H2O 0.75g ；
b. 0.1L of distilled water is added into the millet flour and the peptone, and the mixture is diluted to 0.5L of mixed solution after being uniformly mixed.
c. Adding 0.5L distilled water into semen glycines powder, heating to boil, filtering with four layers of gauze after 20min, and removing semen glycines dregs to obtain filtrate of semen glycines powder.
d. Mixing the liquids obtained in steps b and c, and adding 2g KH2PO4And 0.75g MgSO4·7H2O, uniformly stirring, pouring into a 1L measuring cylinder, and complementing the mixed solution to 1L by using distilled water to obtain a mother solution culture medium;
e. pouring 100mL of mother liquid culture medium into a 250mL triangular flask with a 100mL measuring cylinder, and plugging a silica gel plug at the bottle mouth;
f. sterilizing with high pressure steam of 0.1Mpa at 121 deg.C for 20 min;
g. cooling the mother liquid culture medium, and sterilizing with ultraviolet lamp in a superclean bench for 30 min;
h. under the aseptic condition, a 0.5cm multiplied by 0.5cm agaricus bisporus W192 bacterial block on the PDA inclined plane is inoculated into a mother liquor culture medium, the culture temperature is 25 ℃, and the fermentation culture is carried out for 8d by a 180r/min rotary oscillator, thus completing the preparation of W192 mother liquor strains.
2. Preparation of liquid stock culture medium of W192 fermentation tank
After preparing the W192 mother liquor strain, preparing a W192 fermentation tank liquid stock culture medium by the following method:
a. weighing 30g of millet powder, 12g of soybean powder, 3g of yeast powder and KH according to per liter of liquid stock culture medium2PO4 1.25g，MgSO4·7H2O,0.75g。
b. Accurately weighing semen Setariae powder, semen glycines powder, Yeast powder, KH powder according to 40L liquid stock formula2PO4And MgSO4·7H2And O, adding 4L of water, mixing uniformly to obtain a culture medium concentrated solution, pouring into a fermentation tank with the capacity of 100L, adding 20mL of foam, adding water for diluting until 40L of culture medium is heated to 80 ℃, starting an air pump for ventilation, pouring the prepared culture medium concentrated solution into the fermentation tank, adding the foam, and sealing the fermentation tank.
c. Sterilizing the culture solution at 121 deg.C under 0.1Mpa for 20min, and cooling to 24 deg.C;
d. under the protection of flame, 400mL of liquid seeds are inoculated into a fermentation tank liquid stock culture medium;
e. after inoculation, the ventilation rate is 1.5m3And/min, performing fermentation culture for 7d, and completing the preparation of the W192 fermentation tank liquid stock.
3. W192 respiration bag cultivar production
And (3) performing aseptic operation in a hundred-grade clean space, inoculating the W192 fermentation tank liquid stock into a culture medium of the breathing bag cultivated species, and culturing at 25 ℃ for 15 days, wherein the yield can reach 99%.
Example 2:
1. preparation of W192 mother liquor Strain 1L
a. Respectively weighing 25g of corn flour and 10g of soybean accuratelyPowder, peptone 2g, glucose 10g, KH2PO4 1.75g ，MgSO4·7H2O 0.75g ；
b. 0.1L of distilled water is added into the corn flour and the peptone, and the corn flour and the peptone are diluted to 0.5L of mixed liquor after being uniformly mixed.
c. Adding 0.5L distilled water into semen glycines powder, heating to boil, filtering with four layers of gauze after 20min, and removing semen glycines dregs to obtain filtrate of semen glycines powder.
d. Mixing the liquids obtained in steps b and c, and adding 1.75g KH2PO4And 0.75g MgSO4·7H2O, uniformly stirring, pouring into a 1L measuring cylinder, and complementing the mixed solution to 1L by using distilled water to obtain a mother solution culture medium;
e. pouring 100mL of mother liquid culture medium into a 250mL triangular flask with a 100mL measuring cylinder, and plugging a silica gel plug at the bottle mouth;
f. sterilizing with high pressure steam of 0.1Mpa at 121 deg.C for 20 min;
g. cooling the mother liquid culture medium, and sterilizing with ultraviolet lamp in a superclean bench for 30 min;
h. under the aseptic condition, a 0.5cm multiplied by 0.5cm agaricus bisporus W192 bacterial block on the PDA inclined plane is inoculated into a mother liquor culture medium, the culture temperature is 25 ℃, and the fermentation culture is carried out for 7d by a 160r/min rotary oscillator, thus completing the preparation of W192 mother liquor strains.
2. Preparation of liquid stock culture medium of W192 fermentation tank
After preparing the W192 mother liquor strain, preparing a W192 fermentation tank liquid stock culture medium by the following method:
a. weighing corn flour 30g, soybean flour 12g, peptone 3g, KH according to per liter of liquid stock culture medium2PO4 1.25g，MgSO4·7H2O,0.75g。
b. Accurately weighing corn flour, soybean flour, peptone and KH according to 40L liquid stock formula2PO4And MgSO4·7H2And O, adding 4L of water, mixing uniformly to obtain a 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 foam enemy, adding water to dilute the mixture until the water temperature in the fermentation tank reaches 80 ℃, starting an air pump to ventilate the mixture, and pouring the prepared culture medium concentrated solutionAdding into fermentation tank, adding DIDE, and sealing the fermentation tank.
c. Sterilizing the culture solution at 121 deg.C under 0.1Mpa for 20min, and cooling to 24 deg.C;
d. under the protection of flame, 400mL of liquid seeds are inoculated into a fermentation tank liquid stock culture medium;
e. after inoculation, the ventilation rate is 1.5m3And/min, performing fermentation culture for 7d, and completing the preparation of the W192 fermentation tank liquid stock.
3. W192 respiration bag cultivar production
And (3) performing aseptic operation in a hundred-grade clean space, inoculating the W192 fermentation tank liquid stock into a culture medium of the breathing bag cultivated species, and culturing at 25 ℃ for 15 days, wherein the yield can reach 99%.
Claims (4)
1. A submerged culture method of a fermentation tank for a W192 agaricus bisporus liquid stock seed is characterized by comprising the following steps:
1) w192 mother liquor strain preparation
Cutting the activated agaricus bisporus plate mother seeds into bacterium blocks with the diameter of 1.0 cm by using a hole puncher, transferring the bacterium blocks to 100mL of sterile shake flask seed culture medium in a 250mL triangular flask, and inoculating 2 bacterium blocks to each triangular flask; then rotationally oscillating and culturing for 8d on a culture oscillator with the culture temperature of 24 ℃ and the oscillation rotation number of 180r/min to obtain the agaricus bisporus mother liquor strain;
2) submerged culture of liquid protospecies in fermentation tank
Winding gauze with an iron wire ring with a handle, and pouring 95% alcohol; completely sleeving the flame ring on the inoculation port, opening the air release valve, lighting the flame ring, opening the inoculation port, closing the air release valve of the tank body when the inoculation port is opened to a half, quickly, stably and lightly inoculating strains after the inoculation port is completely opened, and screwing the inoculation port; opening an air release valve of the tank body, adjusting the pressure to be more than 0 and less than 0.05MPa, and entering a culture stage; and carrying out submerged fermentation culture for 7d to obtain an agaricus bisporus liquid stock.
2. The culture method according to claim 1, wherein the W192 mother liquor medium is prepared by a method comprising the steps of:
accurately weighing 7.5g of millet flour and 5.0g of soybean flour, placing in a stainless steel pot, adding 500mL of water, boiling for 5min, adding 10g of glucose, 2g of peptone and MgSO 24·7H2O 0.75g，KH2PO42.0 g of the seed culture medium is uniformly stirred by a glass rod, the volume is fixed to 1000mL by using a measuring cylinder to obtain the shake flask seed culture medium, the prepared culture medium is placed on a magnetic stirrer to be subpackaged, each triangular bottle contains 100mL, the culture medium is placed in a sterilizer to be sterilized at 121 ℃ for 30min, and the sterilized culture medium is naturally cooled for later use.
3. The cultivation process as claimed in claim 1, wherein the preparation of the fermenter liquid seed culture medium comprises the following steps:
(1) weighing corn flour or millet flour 30g, soybean flour 12g, yeast powder 3g and KH per liter of liquid stock culture medium2PO41.25g，MgSO4·7H2O,0.75g is prepared;
(2) weighing a fermentation tank liquid stock culture medium according to a formula, placing the fermentation tank liquid stock culture medium 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, starting air pump to ventilate when water temperature in the fermentation tank reaches 80 deg.C, pouring the prepared culture medium concentrated solution into the fermentation tank, adding DIDE, sealing the fermentation tank, sterilizing at 121 deg.C for 30min, and cooling to obtain fermentation tank liquid stock culture medium.
4. The cultivation process as claimed in claim 1, wherein the sterilization process of the fermenter liquid seed culture medium comprises the following steps:
1) water is added into the interlayer of the tank body
Firstly, adding water into an interlayer of a liquid fermentation tank, wherein the method comprises the following steps: the silicone tube is connected with the water pipe and the interlayer water inlet valve, and the interlayer water outlet valve is opened to add water so as to prevent the interlayer pressure from being overhigh and damaging the inner tank body and prevent the water from being added to the interlayer water outlet valve to output water;
2) liquid culture medium sterilization
Adjusting the temperature of the temperature control box to 121 ℃, opening the heating rod to start heating, opening the interlayer water drain valve, and draining the virtual pressure and excess water in the interlayer;
2.1) adding Water to the tank
Adding water into the tank body through the inoculation valve or the inoculation port until the temperature of the water rises to 60 ℃;
2.2) addition of the 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, adding the liquid height to the tank body observation mirror, adding 20mL of defoaming agent, screwing down a fermentation tank cover, opening an air pump, and opening the air pump: opening a filter shell air release valve, opening a filter shell air inlet valve, then 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: closing the air pump, closing the air release valve and the air inlet valve of the filter shell, and then closing the air release valve of the tank body; continuously heating;
3.3) Sterilization of the liquid Medium
Closing the interlayer water discharge valve after hot steam is discharged for 3-5 min, closing the interlayer air outlet valve, opening the interlayer air outlet valve and then opening the tank air inlet valve when the interlayer pressure is increased to 0.1MPa, opening the tank air outlet valve slightly, and closing the tank air outlet valve after the air inlet pipe burns hands; when the pressure gauge of the tank body reaches 0.15MPa, timing is started, a tank body air release valve 1/3 is opened, and the temperature is adjusted according to the pressure to keep for 30min for sterilization;
3.4) temperature reduction
After the timing is finished, closing the air inlet valve of the tank body, closing the air outlet valve of the interlayer, adjusting the temperature of the temperature control meter to 25 ℃, and closing the heating rod;
3.5) placing interlayer hot water
Opening a large air release valve, and releasing hot water in the interlayer through an interlayer water inlet valve until the pressure of an interlayer pressure gauge is reduced to 0;
3.6) liquid Medium Cooling
Opening an interlayer water drain valve to connect a water drain pipe to a sewer, and cooling the interlayer water inlet valve through a silica gel pipe to be connected with a water inlet pipe; when the pressure of the pressure gauge of the tank body is reduced to 0.05MPa, the air pump is started, the tank body is prevented from generating negative pressure in the cooling process to cause pollution, the lower cold water is cooled upwards quickly, and the air pump is started: opening the air release valve of the filter shell, opening the air inlet valve of the filter shell, then opening the air pump, closing the air release valve of the filter shell, discharging the condensed water through the small air release valve of the filter shell in the culture process, adjusting the pressure of the tank body to be 0 MPa-0.05 MPa through the air release valve of the tank body, and starting inoculation when the temperature is reduced to 24 ℃.
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Citations (4)
| 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 |
| US20090235579A1 (en) * | 2006-05-25 | 2009-09-24 | Cj Corp. | Method of culturing agaricus bisporus mycelium and medium for culturing the same |
| 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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Patent Citations (4)
| 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 |
| US20090235579A1 (en) * | 2006-05-25 | 2009-09-24 | Cj Corp. | Method of culturing agaricus bisporus mycelium and medium for culturing the same |
| 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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| CN112655463B (en) | 2023-08-11 |
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