(1) Large-scale cultivation of Lentinus edodes is arranged in the production schedule, and there are a large number of strains. The required strains should be cultivated before the production barrel (bag), and the age of the strains is 60 ~ 90 days. If the age of bacteria is too short or too long, the survival rate of colonization after inoculation is slow, which is not conducive to improving the yield of bacteria tubes (bags). In particular, the over-summer strain has low survival rate and is easy to be infected with miscellaneous bacteria, so it should be eliminated. The specific date of strain production should be determined according to the production progress of gas cylinders. (2) Dosage of strains The dosage of strains is mainly determined by the cultivation scale and inoculation density. According to 4-6 holes per bag, 70-80 bags of strains can be inoculated per kilogram. Mushroom farmers should order strains from reputable strain factories in advance according to the cultivation plan, and put forward the main variety names, strain quantity and quality requirements. It is not appropriate to buy everywhere for the time being, otherwise it will easily fail.
The production of strains must comply with the relevant laws and regulations of the state. Conditional units or individuals can refer to the following methods for mushroom strain production. Those engaged in the breeding of edible fungi varieties and the production, operation, use and management of strains shall meet the requirements of the Measures for the Administration of Edible Fungi and the Technical Regulations for the Production of Edible Fungi (NY/T 528). Please refer to the relevant materials for the technological process and technical points of mushroom strain production, and this book will not introduce them in detail.
Several problems should be paid attention to in the production and use of Lentinus edodes strains: (1) Obtaining pure mycelium culture by tissue separation ① Selecting seed mushrooms. As seed mushrooms for tissue isolation or spore isolation, it is necessary to select seed mushrooms with early fruiting, round shape, short stalk, thick meat, moderate size, no pests and diseases, and mature within 5-6 minutes (partial rupture of the inner membrane of the mushroom) from high-yield populations with strong stress resistance as seed mushrooms. Because the first and second batches of mushrooms are more precocious, the mushroom sticks are rich in nutrition at this time, and the produced mushrooms are robust. It is easy to obtain pure mycelium culture (strain) with the characteristics of this variety (strain) by spore separation or tissue separation. Therefore, it is suggested to select seed mushrooms from the first and second batches of mushrooms. This principle is applicable to a variety of edible mushrooms, such as mushrooms, straw mushrooms, Pleurotus ostreatus, Pleurotus nebrodensis, Pleurotus eryngii, Pleurotus ostreatus, Flammulina velutipes and so on. ② Tissue separation. Select seed mushrooms according to the above standards, and put them into sterile paper bags after harvesting (plastic bags are not allowed). Under aseptic conditions, firstly, the surface of the mushroom was disinfected with alcohol cotton balls, and then the mushroom was torn in half by holding the stalk. After taking the scalpel for flame sterilization and cooling, pick a small piece of mushroom meat (the size of rice grain) at the junction of mushroom cover and mushroom handle, transplant it on the slope of PDA (prepare it in advance and sterilize it for later use), and then turn to the culture observation stage. ③ Cultural observation. After 3 ~ 5 days at 25℃, white fluffy hyphae grew on the surface of tissue isolate and grew in a star shape on the culture medium. During the culture period, check 1 time every 1 ~ 2 days, and eliminate the cultures contaminated by mold or bacteria at any time. After 10 ~ 15 days of culture and 1 test tube culture, the mother strain of Lentinus edodes can be obtained. The cultivation experiment was carried out to confirm that it can produce mushrooms normally before it can be used for strain production and cultivation. The author found that many strain producers and individual mushroom farmers isolated Lentinus edodes strains themselves, and cultivated them directly with tissue isolates of Lentinus edodes without conducting cultivation experiments at all. Production practice shows that this is very dangerous and the loss is often unexpected. (2) There is a strict technical route between strain production and strain production by using edible fungi, which can only be done step by step. In the production of Lentinus edodes strains, the pure mycelium culture and its transferred strains obtained by the separation of fruiting body tissue, spores or mushroom substrate mycelium are usually called mother strains, also known as first-class strains. The pure culture of mycelium expanded by mother seed transplantation is called original seed (sometimes called mother seed) and also called secondary seed. The original seed is mainly used to produce cultivated seeds, and can also be directly used as cultivated seeds. The pure culture of mycelium transplanted and expanded from original seeds is called cultivated seeds, or production seeds, also known as tertiary seeds. Cultivated species are directly used for cultivation.
Inspection of strain quality: The inspection of mushroom strain quality can refer to the national standard GB 19 170-2003. Check the quality of the original and cultivated strains according to the following steps: (1) Check the seed production records to see if they are worthy of the name, so as to avoid arrogance. (2) Calculate the yield of the strain. Only strains with low pollution rate can be used as breeding and sowing strains. Contamination of miscellaneous bacteria often occurs when making strains. During the culture period, it is normal that the impurity pollution rate of the original seed is ≤5%, or the impurity pollution rate of the cultivated seed is ≤ 10%, and the impurity-removed strain can be used as planned. If the pollution rate of the original species is > 10% and that of the cultivated species is > 20%, it means that the pollution rate is too high. Although no miscellaneous bacteria were found in the remaining strains after careful inspection, it is difficult to determine that they are indeed pure cultures (strains) without miscellaneous bacteria. The production practice shows that when the pollution rate of the original species is too high, the remaining bacteria-free strains should be replanted and used directly as cultivated species for production.
If the contamination rate of the original seed exceeds 15% and the contamination rate of the cultivated seed exceeds 25%, it is not suitable for strain production and can be directly used to cultivate mushrooms or ears; Or all of them are discarded, and after sterilization, the culture material is taken out for alternative cultivation of edible fungi. (3) Investigate the culture time of the strain. Under suitable culture conditions, the strain can be completely eaten after about 25 days of culture. If the culture time is too long, and the feed has not been completely eaten for more than 45 days, it means that the strain is aging and not suitable for production. (4) Regeneration test In order to avoid purchasing or using weak, degraded or aging strains, a regeneration test should be conducted before purchasing or using to check whether the strains are robust. The specific method is as follows: take a piece of soybean or broad bean-sized strain among the strains and transplant it into a Petri dish. First, spread a piece of filter paper or several layers of toilet paper in the Petri dish, add a little cold boiled water to moisten the paper, and cultivate it at room temperature (25℃) for 3-5 days. If the seed block germinates normally and the mycelium grows vigorously and orderly, it shows that the strain is robust and of good quality. If there is no change after 5 ~ 7 days of culture, it shows that the strain has weak growth and poor regeneration ability, so it is not suitable for utilization.
In the process of culture, storage and transportation, strains with damaged containers or visited by rats and pests shall not be used. (5) Check the maturity of the strain. In production, the survival rate of inoculation can be improved by cultivating mature strains. Judging whether the strain is mature or aging can generally be observed and analyzed from three aspects:
① Observe whether the strain is lumpy. Whether it is sawdust or wood, part of the mycelium enters the culture medium and part extends into the air, which is called aerial mycelium. The aerial hyphae of healthy strains are very vigorous. When stretched to a certain extent, they connect with each other, wrapping the culture medium into blocks and turning it into quite tough blocks, which is the sign of a good strain.
② Observe whether the surface of the strain is normal. After the mushroom was cultured for too long, the epidermis gradually became thick and brown, similar to the color of the mushroom lid surface. When this color appears on the surface of the strain, it means that the strain has aged. Aging strains have weak regeneration ability and are easy to be infected with miscellaneous bacteria. If miscellaneous bacteria pollution is found, it will be eliminated in time. The aging phenomenon of wood block bacteria is due to the excessive corruption of mycelium on wood, and the cork tissue is softened, which is easy to be crushed. Like immature strains, the mycelium of this kind of aging strains is not easy to penetrate into wood after inoculation, and the colonization survival rate is low, so it is not suitable for utilization.
③ Observe whether the discoloration of the culture medium is uniform. Lentinus edodes is a kind of white rot wood rot fungus, and the wood decays into yellowish white after mycelium utilization. Uniformity of medium discoloration can be used to judge whether the strain is mature. When checking sawdust strains, you can take out several bottles at random, and take one piece from the middle of each bottle to observe whether the discoloration of sawdust is uniform and the degree of discoloration, whether hyphae only spread to the outer layer and whether there are local aseptic spots, and then judge the maturity of the whole batch of strains through samples. If it is not mature, we must continue to cultivate. Through the above identification, qualified strains are marked for use. If you don't inoculate in the near future, you must transfer the strains to the cold storage or store them in other clean, cool (15℃ below), dry and well-ventilated places to avoid stacking with pesticides and fertilizers, so as not to affect the mycelium vitality.
Other problems that should be paid attention to in the process of making and using strains: (1) It is not suitable to transfer edible fungi strains for many times. Edible fungi strains are pure mycelium cultures. In production, the mother seed can not be directly used for sowing, and it must be expanded and cultivated into original seed, cultivated seed or liquid strain before it can be widely used in production. Most of the mother seeds purchased from scientific research institutions have been transferred for 2 ~ 3 times, and it is safer to transfer tubes 1 time in the grass-roots strain factory. If too many tubes are changed in the spawn factory, there will often be a phenomenon of slow mycelium growth, or delayed or less fruiting, or small fruiting with poor quality, or even a phenomenon of not harvesting mushrooms. All of the above are called strain degradation. Therefore, it is not advisable to change the tube many times.
It is an accepted fact that repeated tube transfer will lead to strain degradation. If the reason is investigated, there is still no conclusion. However, the consistent view is that high temperature mutation and virus parasitism lead to strain degradation. Of course, repeated tube rotation is not the only reason for the degradation of the strain, and the so-called Buller phenomenon is also one of the reasons for the degradation of the strain. (2) Standards of excellent strains All kinds of edible fungi strains have special and identical quality standards. These same quality standards are mainly pure and powerful.
① First, the genetic germplasm of the strain is excellent. Production practice has proved that it can achieve the goal of high quality and high efficiency. This kind of strain is the Lentinus edodes strain with excellent genetic germplasm, which is the so-called main cultivated variety.
② The second strain is pure strain. Except for tremella species, other cultivated edible fungi are pure mycelium cultures, and other strains other than mixed bacteria or pure cultures are not allowed. For example, mushroom strain 856 can only be a pure culture of mycelium of strain 856. First, do not mix any mold, bacteria or virus such as Mucor, Rhizopus, Aspergillus and Trichoderma; Second, the mycelium of Pleurotus ostreatus, Flammulina velutipes and other edible fungi should not be mixed; Thirdly, it is not allowed to mix any Lentinus edodes except Lentinus edodes strain 856, such as Wuxiang 1, 135- 1, 939, etc.
③ The third is the virulent strain. The strain has normal appearance, vigorous growth and strong regeneration ability after inoculation (transplantation), which is called strong. The strain is strong and can grow and reproduce normally as soon as possible after inoculation. If the strain is not strong enough to germinate after inoculation, even if it is not polluted for a long time, it will inevitably affect the normal fruiting.