(A) The mother mushroom and the original species are separately separated and purified from the cultured white fungus mycelium and incense mycelium in a test tube. When producing mothers, first remove the white fungus mycelia and expand it to the required number of tubes. Then cultured at 25 °C to approximately 1cm diameter of Tremella. At this point, in each test tube with long mycelium growth, inoculation of a small amount of ash mycelium, when the two are together, they can be sold or used for the production of the original species.
If the ash is used first, the mycelium of the ash grows quickly and quickly covers the slope and quickly consumes nourishment. After that, the white fungus mycelium attached to it is not easy to grow.
If the cotyledon hyphae and tremella mycelia are inoculated in a test tube at the same time, there will also be defects of less mycelium of the white fungus and a large number of mycelia.
Therefore, the production of Tremella parent species (test tube species) must pay attention to the order of inoculation, and pay attention to control the ratio of the two.
(II) Tremella and its "associated bacteria" In the food and medicine that can be cultivated artificially, only the white fungus and golden ear are present in this type of bacteria. The formation of their fruit bodies, Tremella in addition to the need to have pure white fungus mycelium, but also must have the "associated bacteria" that is, the help of incense gray bacteria; gold ear in addition to pure gold ear mycelium, but also must have the help of hairy hard leather Both are indispensable. Under normal conditions, the pure mycelium of Tremella fuciformis and Auricula grows very slowly, and its ability to decompose lignin and cellulose is very weak. It depends on the decomposition of the substrate by the hyphae and the coarse hairs and harder hyphae to provide their growth and development. Nutrients, otherwise it is impossible to form fruiting bodies. Therefore, in the propagation of Tremella fusca or Golden ear, it is necessary to transfer the two corresponding mycelia onto the same medium for mixed culture. The tremella is introduced as an example to introduce two kinds of mycelium mixed seed production technologies:
1. Inclined female parent hybrid method (Figure 33-1) Take the Tremella mycelium test tube mother and new
Each PDA bevel test tube is placed in the inoculation room (box) along with the inoculation device, and transfer is performed according to the asepsis operation, ie, picking up the size of the rice grains, cultivating a piece of the culture medium, and cultivating the slant surface of the blank test tube. On the base, add tampons and incubate at a constant temperature of 22-24°C for 6-8 days. When the white fungus mycelium grows to a size similar to an embroidered ball, such as soybeans, it is transferred to the inoculation room (box) together with the gray ash seedlings and inserted into the inoculation chamber (according to aseptically used inoculum shovel) at a distance of about 2 cm from the tremella mycelia. A small piece of the mycelium of the mycelium was used as the mother of mycelium. The white mycelium was observed on the inoculation block after 2 days of culture. The white hairball was formed around 7 days and covered the pure mycelium of white fungus. It was seen on the top of the white hairs in 12-15 days. Red and yellow beads are successful.
It is necessary to point out that when using the mixed mother of Tremella and Coprinus comatus to expand the original species, the front and rear ends of the incline should be cut off, and only the white fungus mycelium in the form of hydrangea and the black grey mycelium near it should be used for transfer. . Therefore, only one bottle of original species can be connected to one beveled mother species.
Inclined parent species mixing method
Bottled sawdust original mixed method (Figure 33-2) If there is no Tremella pure hyphae, the spore mother of spores cultured spores, and then access to bottled wood culture medium made of Coprinus species, mixed culture.
According to the inoculation requirement, the ratio of 1-15 incense ash beveled mother-species to 12-15 bottles of sawdust medium is used to place the Coprinus comatus mother tube and the sterilized bottled medium and the inoculation utensils together. In the box, aseptically, use a inoculation shovel to pick out the mother of Coprinus comatus with the size of soybean and insert it into the sawdust medium bottle. After the inoculation was completed, the culture was moved to 24-26°C and the germinated white hyphae were observed on the inoculation block after about 2 days. Then check the next day and found that the bacteria were detected and removed in time. Generally about 11 days of culture, the hyphae mycelium can extend about 3 cm into the culture medium. At this time, the surface of the culture medium can be observed to have taupe or dark brown secretions. This indicates that the hyphae grow vigorously and should be inserted in time. Spore mother.
According to the ratio of 1:12-15, take the Tremella sporum mother test tube and the culture medium of cultivated woody culms from Coprinus comatus, and put them into the inoculation room (box). Aseptically, use a sterile inoculating knife to cut the incense On the surface of the mycelium, a small amount of Tremella spores was picked up with a sterile inoculation spatula and inserted into the mycelia of the incense (Figure 33-2). After mixing, the bottles were transferred to culture at 23-25°C, and thick white hyphae could be seen in the spores of Tremella sp. at about 10 days. After about 18 days of cultivation, red and yellow water droplets appeared. The kinks of the twisted fruiting bodies were observed around 25 days, indicating successful culture. One bottle of this mixed wood mixture can be expanded to 30-40 bottles of the original species (secondary species).
Bottled wood chips original mixed method
1. Spores of Tremella fuciformis; 2. Corygium sp.
4. Pick the spores of white fungus; 5. Access the bottle of incense bacteria
(3) The production of planting species depends on the growth of the original species, the size of the ear base, and the depth of the "eating material" of the white fungus mycelium to determine the number of bottles to be expanded per bottle. In order to ensure quality, try to control within 40-60 bottles. If the original species can be stirred very evenly, the mycelia of the white fungus and the mycelium of the ash will be mixed very evenly, and each bottle of the original species may also be connected to 100-200 bottles.
(4) Precautions To produce excellent Tremella species, in addition to the above-mentioned requirements, such as aseptic operations, attention must also be paid to the following aspects:
1. The specific combination of bio-specific white fungus mycelium and ash hyphae also has specificity. Therefore, in order to produce effective Tremella species, we must obtain the pure culture of Tremella (including yeast-like conidia or Tremella mycelium) and the pure ash hyphae respectively from the same ear. The white fungus mycelia and incense hyphae mycelium obtained from different ears or trees in different regions or in the same area cannot be exchanged and combined randomly. Otherwise, there is the possibility of decline. The effect of the exchange and combination must be based on the conditions of the colonization of Tremella mycelia in the wood pulp medium, the size of the ear base, and the condition of opening. Any tremella mycelium that can continue to grow, form an ear base, and open smoothly, can continue to expand and be used for production, otherwise it cannot be used.
Inoculation ratio: According to the slow growth of Tremella mycelia, the growth of Cotyledon mycelium is very fast
This feature, in the pure mixed culture, must pay attention to the ratio of inoculation of Tremella mycelia and incense mycelium. In order to achieve the control of the proportion of both inoculation, using the first culture of tremella mycelium, followed by the method of the incense hyphae, the most ideal. If the inoculation ratio of the white fungus mycelium should reach 1000:1, it is not excessive. (The Tremella mycelium develops to a diameter of about 1 cm. There are tens of thousands of mycelium in it, so long as some hyphae are added )). Otherwise, the white fungus mycelium is rare, and the incense hyphae account for absolute advantage. In the future, the ears will not be eared or the ear rate will be very low. If strains are produced by adding yeast-like conidia to the coniferous mycelium, the number of conidia in the tremella species is large, and the number of conidia in the trachoma yeast-like culture medium per ml may be sufficient. Up to 1 billion to 10 billion. A yeast-like conidia solution is added to the woodchip medium on which the mycelium grows, and the yeast-like conidia are dispersed in the wood pellets, and not all the yeast-like conidia can germinate into hyphae. Therefore, using this method to produce Tremella species, the effect is not very satisfactory, if the Tremella mycelia can not be obtained, the conidia of the Tremella fusca can be concentrated on the surface of the wood culture media of the long ash mycelium, so that the Tremella yeast shape Conidia are easier to germinate.
3. Cultivation condition Tremella pure species (yeast-like conidia or Tremella mycelium)
There are some differences between the culture conditions and the culture conditions of Tremella species and planting species (mixed culture). Therefore, we must pay attention to the conditions of mixed culture, so that the white fungus mycelium and incense mycelium can adapt to the same medium, the same culture conditions. For example, the temperature is controlled in the range of 20-28C (preferably 23-25C). The moisture content of the original species is controlled at 65%, and the moisture content of the planting species is controlled between 65% and 70%. Because the original kind of medium is a little dry, it is conducive to the growth of white fungus mycelium, and it is not easy to produce yeast-like conidia, which is conducive to expanding into planting species; and planting species are a little wet, which is conducive to the production and reproduction of yeast-like conidia. The spore fluid infiltrates deep into the sawdust medium and is evenly mixed with the incense ash hyphae, which can make up for the defect that the "treasure" of the Tremella mycelia is not deep and only grows on the surface, so that the ear rate is higher and more stable.
It is also necessary to determine whether the original species can be expanded into planting species based on the fertility status of each bottle of the original species, and to expand the number of bottle types for each bottle. After mixed culture, the conidia of Tremella fuciformis can be germinated, the mycelia of Tremella fuciformis can continue to grow, and the combination of the ear base and the ear base can grow continuously. All of them are useful species. The original species has been carefully selected, where the ear group is large, and the white fungus mycelium grows more (deep in the material), and the ash gray mycelium is thick and strong, and the number of bottles expanded after stirring is more. On the contrary, it should not be expanded into planting species. For better safety, it is advisable to increase 40-60 bottles per bottle. In conclusion, each bottle of freshly inoculated medium must have tremella mycelia and incense hyphae, or yeast-like conidia and incense hyphae. Or, after the original species is stirred, the cotton plug is stuffed and cultured for another 3-5 days to regenerate the new mycelium of the dismantled white fungus mycelium, so as to ensure the quality of the planted species.
4. Purification and rejuvenation to prevent bacterial species due to long-term use, decreased viability, ear yellow, difficult to open the film, poor regeneration of the disease. According to the production method, different ages of bacteria and different physiological maturity of Tremella fusca strains were selected. Such as Duanmu planting, should be selected small age, low maturity, white fungus aerial mycelium - white hair group is strong, not easily gelatinous strains; bottle planting, should be selected bacteria Lingda, high maturity, white hair Small, easy to gelatinize and easy to open the strains. Aspergillus mycelium should be selected as a strain that has strong wall climbing power, long feather-like branching, and strong wood decomposition ability as a strain for planting wood, but when the material is planted, the incense ash should be slightly weakened by some strains. Otherwise, the full bottle and the full bag are incense ash mycelium, limited nutrients are consumed by the incense gray mycelium, white fungus does not grow.
The original species was stored for too long, the ear was very large, the nutrient consumption of the culture medium was excessive, and the viability of the strains had declined. They should be discarded. Otherwise, the use of the effect is not as good as the cultivation of seedlings of the right age and the expansion of cultured strains.
5. The female parent species should be stored in a refrigerator at 4-15°C. Transplant once every 1-3 months. If the original species is not used immediately, it should also be stored in a cool, dry place. Planting the bottle and planting it within 45 days should be planted as soon as possible.
The pure fungus Tremella (Tremella mycelia and yeast-like conidia commonly known as spores) was preserved separately for a long period of time. The long fruiting bodies had a tendency of yellowing. The mixed cultures were obtained after being preserved for a long period of time. Not easy to yellow. In order to obtain the white fungus, the species should be selected from white and white fungus (wild or domestic). Protection of color should be more than 3 years.
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