Black fungus seed production technology
2020-10-08 16:14:16
(a) Preparation of parent species
The slant culture medium is made of test tubes, and the original test tube seed is inoculated (retained species, and the parent species used for production is the production mother species, which can be used to expand the original species.
1. Medium formula
Formula One Integrated Potato Culture Medium
Potato 200 g Glucose 20 g Potassium hydrogen phosphate 2 g Magnesium sulfate 0.5 g
Vitamin B 110 mg agar 20 g water 1000 ml PH Natural
Formulation II Synthesis Medium
Potato 20 g Glucose 20 g Potassium dihydrogen phosphate 2 g Peptone 10 g Beef paste 5 g
Yeast extract 5 g magnesium sulfate 0.8 g water 1000 ml PH natural
2. Medium production method
Wash the potatoes, peel and cut them into a book, weigh 200g, add 1000ml of water, boil for 15-20 minutes, filter with 4 layers of gauze, take the filtrate and make up to 1000ml of water, add 20g of agar and reheat After the agar melts, add the other ingredients in the formula.
The sub-packing of the culture medium should be carried out before the culture is solidified (40°C solidification in agar medium). The funnel is usually used, or the bottle is divided into bottles. The lower end is connected to a flexible rubber tube, and the lower section of the rubber tube is connected to a water-section glass tube. Install a water stop spring clip. When dispensing, insert the fine glass tube into the tube, but do not touch the tube wall. Each tube should be loaded with 1/4 of the tube length. After the tampon is stuffed, tie a bundle into each 10 test tubes. The tampon is sealed with plastic paper and tied with a string.
3. Sterilization
The bundled test tubes are bundled and placed in a high-pressure steam sterilizer. The bacteria are sterilized under a pressure of 1 kg/cm (pot temperature of 121° C.) for 20 minutes to kill all microorganisms including bud color. When the pressure drops to 0 degrees, the gas in the pan is slowly released. When the medium does not solidify, the test tube is tilted.
4. Inoculation, culture
Aseptically insert the strain into the slant of the test tube, set the temperature at 28°C for 3 days, continue the incubation at 24°C, and continue for 12-15 days. The hyphae overwhelm the entire test tube slant. .
(B) The original species, preparation
The parent species is inoculated into the original culture medium, and the expanded breeding is the original species. The original species is generally used in wide mouth bottles or bags.
Formulated wood chips bran medium
Broad Trees 78% Bran 20% Sugar 1% Gypsum Powder 1% Water 65% PH Natural
Formula II cottonseed husk mixing media
Cotton seed hull 90% sugar 1% bran 5% superphosphate 3% gypsum powder 1% PH value natural
2. Mixing and bagging
Fresh and mildew-free raw materials are selected. Calculate the total amount of ingredients according to the quantity needed for production. Bran and gypsum powder are uniformly mixed in the culture medium. After sugar and superphosphate are dissolved in water, water is added according to the amount of dry materials mentioned above. , The ratio is 1:1, 3 mix well, the moisture content of the culture material is appropriate 60%, the water content is suitable for the water dripping between the fingertips of the training material. After 30 minutes of boring, bottling (bags). Slightly pressurize with mechanical or manual bagging. (The plastic bag size is usually 9*21*0, 04, and the bag width is 9cm, length is 21cm, and the single layer is 4cm thick.) In fact, a plastic ring is put on the mouth of the bag to form a bottle, and then it is in the mouth of the bag. Plugs are suitable for tampon or kraft wrap.
There are two kinds of original sterilization methods, autoclave and atmospheric sterilization. Autoclave sterilization, also known as pressurized steam sterilization, sterilization principle: In the high temperature (120 °C), high pressure (1 kg / cm2) sterilization under conditions for 2-3 hours, the spores, including all the microorganisms killed. Atmospheric pressure sterilization refers to circulating steam sterilization. Due to different sterilization equipment, conditions, and basic factors, the temperature changes between 90-103°C. The heat and penetrating power are not strong enough to maintain at a certain temperature for a long time. For bacterial purposes, it is usually maintained at 100°C for 8-10 hours.
4. Inoculation and culture
Regardless of vaccination or vaccination room vaccination, inoculation or inoculation chambers must be sterilized with 0 or 2% lycopene or bleach before inoculation, and then the sterilized bottle (bag) is placed in the vaccination kit. Inside or in the inoculation room, after irradiating with an Ultraviolet Lamp for 30 minutes, inoculation was carried out under aseptic conditions, or airtight disinfection with an aerosol prior to inoculation. When the aerosol is ignited, the doors and windows should be closed. Inoculation room disinfection, a room to be ignited 3 boxes of 50 grams of the size of aerosol; inoculation box disinfection only use a weight of 40 grams each time, but subdivided into 20 packets of which 2 packets can be ignited (2 grams per pack). General 18180 centimeters, the mother tube can receive 4-6 bottles (bags) of the original species. Put the inoculated strainer bottle (bag) into the chamber culture. With a culture chamber temperature of 22-26°C and a relative humidity of 80% or less, the light is weakly scattered light, and good ventilation conditions should be maintained during germination. It takes 30-40 days for the original species to reach the bottle (bag) from inoculation.
(III) Preparation of cultivars
The original species is further expanded by breeding. Cultivated species are generally prepared using plastic bags, specifications 15330,05, sealed with a cotton-free cover. A bottle of original species can be connected to 50 bottles (bags) of cultivars. Culture medium formulation and mixing, bagging, sterilization, inoculation, and bacterium administration are the same as those for the original species.
(d) Identification of strains
The quality of strains is not only related to the success of bacteria, but also related to the yield and benefits. The cultivation of fungus must be strictly controlled. The quality identification of strains has two meanings: The first is whether the cultured strains are required for cultivation. The second is whether the strain is a good strain. The normal mycelia of black fungus strains should be white and vigorous. After 45 days of bacterial culture, there will be chrysanthemum-like or plum-type primordium from top to bottom between the bottles (bags), and the color is bright brown to dark brown. If the bottom of the bottle (bag) is light yellow liquid, or yellow mucus appears in the upper half, the medium shrinks and expired strains should be eliminated.
If the mycelium of mycelia only reaches 1/3-1/2 of the bottle (bag), a pale black colloidal base appears, indicating that the strain is precocious, or that it has expanded too much, or because the light in the culture room is too strong. If the hyphae grows only one corner and does not spread, the medium may be too dry, too wet, or not wet or dry. If the mycelial growth has not stopped before and there is a clear line of inhibition, it is mostly caused by bacteria. In the fungus process, it was found that the bacteria should be detected in time.
The slant culture medium is made of test tubes, and the original test tube seed is inoculated (retained species, and the parent species used for production is the production mother species, which can be used to expand the original species.
1. Medium formula
Formula One Integrated Potato Culture Medium
Potato 200 g Glucose 20 g Potassium hydrogen phosphate 2 g Magnesium sulfate 0.5 g
Vitamin B 110 mg agar 20 g water 1000 ml PH Natural
Formulation II Synthesis Medium
Potato 20 g Glucose 20 g Potassium dihydrogen phosphate 2 g Peptone 10 g Beef paste 5 g
Yeast extract 5 g magnesium sulfate 0.8 g water 1000 ml PH natural
2. Medium production method
Wash the potatoes, peel and cut them into a book, weigh 200g, add 1000ml of water, boil for 15-20 minutes, filter with 4 layers of gauze, take the filtrate and make up to 1000ml of water, add 20g of agar and reheat After the agar melts, add the other ingredients in the formula.
The sub-packing of the culture medium should be carried out before the culture is solidified (40°C solidification in agar medium). The funnel is usually used, or the bottle is divided into bottles. The lower end is connected to a flexible rubber tube, and the lower section of the rubber tube is connected to a water-section glass tube. Install a water stop spring clip. When dispensing, insert the fine glass tube into the tube, but do not touch the tube wall. Each tube should be loaded with 1/4 of the tube length. After the tampon is stuffed, tie a bundle into each 10 test tubes. The tampon is sealed with plastic paper and tied with a string.
3. Sterilization
The bundled test tubes are bundled and placed in a high-pressure steam sterilizer. The bacteria are sterilized under a pressure of 1 kg/cm (pot temperature of 121° C.) for 20 minutes to kill all microorganisms including bud color. When the pressure drops to 0 degrees, the gas in the pan is slowly released. When the medium does not solidify, the test tube is tilted.
4. Inoculation, culture
Aseptically insert the strain into the slant of the test tube, set the temperature at 28°C for 3 days, continue the incubation at 24°C, and continue for 12-15 days. The hyphae overwhelm the entire test tube slant. .
(B) The original species, preparation
The parent species is inoculated into the original culture medium, and the expanded breeding is the original species. The original species is generally used in wide mouth bottles or bags.
Formulated wood chips bran medium
Broad Trees 78% Bran 20% Sugar 1% Gypsum Powder 1% Water 65% PH Natural
Formula II cottonseed husk mixing media
Cotton seed hull 90% sugar 1% bran 5% superphosphate 3% gypsum powder 1% PH value natural
2. Mixing and bagging
Fresh and mildew-free raw materials are selected. Calculate the total amount of ingredients according to the quantity needed for production. Bran and gypsum powder are uniformly mixed in the culture medium. After sugar and superphosphate are dissolved in water, water is added according to the amount of dry materials mentioned above. , The ratio is 1:1, 3 mix well, the moisture content of the culture material is appropriate 60%, the water content is suitable for the water dripping between the fingertips of the training material. After 30 minutes of boring, bottling (bags). Slightly pressurize with mechanical or manual bagging. (The plastic bag size is usually 9*21*0, 04, and the bag width is 9cm, length is 21cm, and the single layer is 4cm thick.) In fact, a plastic ring is put on the mouth of the bag to form a bottle, and then it is in the mouth of the bag. Plugs are suitable for tampon or kraft wrap.
There are two kinds of original sterilization methods, autoclave and atmospheric sterilization. Autoclave sterilization, also known as pressurized steam sterilization, sterilization principle: In the high temperature (120 °C), high pressure (1 kg / cm2) sterilization under conditions for 2-3 hours, the spores, including all the microorganisms killed. Atmospheric pressure sterilization refers to circulating steam sterilization. Due to different sterilization equipment, conditions, and basic factors, the temperature changes between 90-103°C. The heat and penetrating power are not strong enough to maintain at a certain temperature for a long time. For bacterial purposes, it is usually maintained at 100°C for 8-10 hours.
4. Inoculation and culture
Regardless of vaccination or vaccination room vaccination, inoculation or inoculation chambers must be sterilized with 0 or 2% lycopene or bleach before inoculation, and then the sterilized bottle (bag) is placed in the vaccination kit. Inside or in the inoculation room, after irradiating with an Ultraviolet Lamp for 30 minutes, inoculation was carried out under aseptic conditions, or airtight disinfection with an aerosol prior to inoculation. When the aerosol is ignited, the doors and windows should be closed. Inoculation room disinfection, a room to be ignited 3 boxes of 50 grams of the size of aerosol; inoculation box disinfection only use a weight of 40 grams each time, but subdivided into 20 packets of which 2 packets can be ignited (2 grams per pack). General 18180 centimeters, the mother tube can receive 4-6 bottles (bags) of the original species. Put the inoculated strainer bottle (bag) into the chamber culture. With a culture chamber temperature of 22-26°C and a relative humidity of 80% or less, the light is weakly scattered light, and good ventilation conditions should be maintained during germination. It takes 30-40 days for the original species to reach the bottle (bag) from inoculation.
(III) Preparation of cultivars
The original species is further expanded by breeding. Cultivated species are generally prepared using plastic bags, specifications 15330,05, sealed with a cotton-free cover. A bottle of original species can be connected to 50 bottles (bags) of cultivars. Culture medium formulation and mixing, bagging, sterilization, inoculation, and bacterium administration are the same as those for the original species.
(d) Identification of strains
The quality of strains is not only related to the success of bacteria, but also related to the yield and benefits. The cultivation of fungus must be strictly controlled. The quality identification of strains has two meanings: The first is whether the cultured strains are required for cultivation. The second is whether the strain is a good strain. The normal mycelia of black fungus strains should be white and vigorous. After 45 days of bacterial culture, there will be chrysanthemum-like or plum-type primordium from top to bottom between the bottles (bags), and the color is bright brown to dark brown. If the bottom of the bottle (bag) is light yellow liquid, or yellow mucus appears in the upper half, the medium shrinks and expired strains should be eliminated.
If the mycelium of mycelia only reaches 1/3-1/2 of the bottle (bag), a pale black colloidal base appears, indicating that the strain is precocious, or that it has expanded too much, or because the light in the culture room is too strong. If the hyphae grows only one corner and does not spread, the medium may be too dry, too wet, or not wet or dry. If the mycelial growth has not stopped before and there is a clear line of inhibition, it is mostly caused by bacteria. In the fungus process, it was found that the bacteria should be detected in time.
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