Production and breeding technology of Flammulina velutipes

Flammulina velutipes is a vegetable with very high nutritional value, so today I will tell you about the production and breeding technology of Flammulina velutipes:

1. Purchase of Flammulina velutipes strains and main facilities and equipment for production

1.1 Main facilities: Inoculation room, culture room, operation room, storage room, the size of which depends on the production scale.

1.2 Main equipment: Clean bench (or inoculation box), constant temperature box, operating table, swivel chair, high pressure sterilizer, normal pressure sterilizer, steel pot, balance, platform scale, measuring cylinder, glass bottle, test tube, etc.

1.3 Selection of strains. Select high-quality, high-yield, strong stress resistance, stable properties, fast growth, high purity, free of foreign bacteria, diseases and insect pests.

2. Selection and production of culture medium for each strain

2.1 Preparation of mother culture slant medium:

Recipe ① 200-250g potato, 20g agar, 20g sugar, 1000ml water, pH adjusted to 6.5. ② 200-250g potato, 20g agar, 20g glucose, 1000ml water, pH adjusted to 6.5. ③ 200-250g potato, 5g magnesium sulfate, 20g sugar, 100ml vitamin B1 (B2), 2.50g potassium dihydrogen phosphate, 20g agar, 1000ml water, pH adjusted to 6.5.

Preparation: Cut the potatoes into slices and boil them in a steel pot for 30 minutes. Use four layers of gauze to filter the juice, add agar and other ingredients, stir to dissolve, then add sugar, add water to 1000 ml, and transfer to a culture test tube with a capacity of 1/5 of the tube length.

2.2 Preparation of culture medium for original species and cultivated species:

Formula ① Sawdust culture medium: 73% broad-leaved tree chips, 1% sucrose, 2.50% fine rice bran (or wheat bran), 1% calcium carbonate, water-to-material ratio 1:1.20-1.30.

② Cottonseed hull culture medium: cottonseed hull 88%, sucrose 1%, fine sugar (or wheat bran) 10%, calcium carbonate 10%, water-to-material ratio is the same as ①.

③Wheat grain culture medium: wheat grains (or corn grains are also acceptable) 99% calcium carbonate 1%, water-to-material ratio 1:1.20.

Preparation: Weigh all ingredients according to the above ratio and mix them. Dissolve sugar and chemicals in water and add them to the material and mix well. The amount of water used for dissolution is recorded in the water-to-material ratio. Pre-wet cottonseed hulls with water before mixing; soak wheat grains before mixing and cook them in a pot until the grains do not break. Take them out and dry them before mixing with other materials. After all the materials are mixed, bottle them, and the amount is 1/3 of the bottle capacity.

3. Liquid strain preparation

Recipe: ① 3% glucose, 1% corn flour, 2% soybean meal, 0.20% calcium carbonate, 0.10% potassium dihydrogen phosphate, 0.50% yeast powder, 0.05% magnesium sulfate. Dissolve each ingredient in water and mix well to make a suspension. Boil for 30 minutes, take the supernatant and dissolve it, then put it into a 500 ml conical flask containing 10 small glass beads.

Formula ② 20% potato, 0.20% protein, 20% glucose, 0.05% potassium dihydrogen phosphate, 0.05% magnesium sulfate, 0.01% potassium chloride. Extract the filtrate from potatoes in the conventional way, add other ingredients and then fix the solution and put it into the bottle.

Recipe ③: 5% corn flour, 0.50% yeast powder, 4% sucrose, 0.20% calcium carbonate, VB1 mg. The preparation method is the same as recipe ①.

3.1 Sterilization of culture medium and disinfection of facilities and equipment

3.2 Slant medium for mother culture. Liquid culture medium should be sterilized by high pressure at 1.10 kg per 1000 cm3 at 121°C for 30 minutes. Solid culture medium should be placed on a slant immediately after sterilization, and the length of the slant should be 1/2 of the length of the test tube.

3.3 Culture medium for stock and cultivar. For high pressure sterilization, use a pressure of 1.50 kg per thousand centimeters and sterilize at 121°C for 1 hour. For culture medium made from wheat grains, the sterilization time should be appropriately extended. Ordinary pressure steam sterilization can be used, that is, sterilization at 100°C for 8 hours.

3.4 Facility disinfection. The inoculation room, culture room, and operation room should be fumigated with formalin heating or sulfur powder burning. The dosage is 10 ml of formalin per cubic meter of space and 15 g of sulfur powder per cubic meter of space. Use calcium hypochlorite (20-40 g per thousand meters) aqueous solution and 2%-3% Lysol to spray the ground and walls.

3.5 Equipment disinfection. The clean bench and its tools should be soaked and scrubbed with 70% alcohol or 5% carbolic acid and 0.25% chlorhexidine. Before inoculation, turn on the ultraviolet light for sterilization for 15 minutes.

4. Inoculation and cultivation of bacteria

4.1 Inoculation and cultivation of mother culture. When inoculating, the test tube (bottle mouth) is facing downward and close to the flame of the alcohol lamp. Use an inoculation needle to cut the mycelium in the mother culture test tube together with the original culture medium into small pieces of 0.50 cm x 0.50 cm. Transfer them to the slope of a new test tube culture medium, plug it with cotton, and place it in a 23℃ constant temperature box or culture room for 8-10 days. The mycelium will cover the slope.

4.2 Inoculation and cultivation of original seed and cultivated seed. Use an inoculation needle to pick a 1 cm2 bacterial block from the mycelium in the mother seed test tube and inoculate it into the culture medium in the original seed (or cultivated seed) bottle. Also, place the bottle mouth close to the flame of the alcohol lamp and do it quickly. After the connection is completed, plug it with cotton and place it in a 23℃ culture room for cultivation.

4.3 Inoculation and cultivation of liquid strains: Inoculate a 2 square centimeter slant mother strain with vigorous growth into the liquid culture medium after high pressure sterilization, so that the side of the vadose mycelium is suspended on the liquid surface, and culture statically at 23℃ for 2-3 days, and then place it on a compound shake bottle machine (oscillation frequency 80-100 times/minute) for shaking culture for 3-4 days. When the liquid is light yellow, clear and transparent, and has fragrance, move it into the sterilized inoculation room, pour the liquid into the surface of the sterilized culture medium at the flame mouth of the alcohol lamp, the amount is 10-15 ml per bottle, and then place it in a culture room at 23℃ for about 27 days to be used as a culture. The characteristics of liquid strains are that they can be used as original strains when the original strain amount is small. It only takes about 1 week to culture with liquid culture medium, which can be shortened by 4 times compared with the culture of solid original strains. In addition, the age of the bacteria is consistent, the mycelium is healthy, and the vitality is vigorous.

5. Preservation of Flammulina velutipes strains

5.1 Low temperature storage. Place the bacteria that have grown on the slope in a 4°C refrigerator and transplant them once every 5 months.

5.2 Store at room temperature. After the mycelium of the culture medium prepared with sawdust is fully grown, it should be sealed with paraffin and cotton plugs and stored at room temperature.

5.3 Liquid nitrogen storage. The strains are stored in liquid nitrogen tanks at -130-- -193℃.