A method for obtaining spore powder by culturing ganoderma lucidum on linden wood with solid-liquefied bacteria
By using a special solid-liquid culture medium for Ganoderma lucidum cultivation on linden wood and a two-stage solid-liquid culture method, the problems of low production efficiency and contamination risk of Ganoderma lucidum spore powder have been solved, achieving efficient and stable spore powder production.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- ZHEJIANG UNIV
- Filing Date
- 2026-02-28
- Publication Date
- 2026-06-23
AI Technical Summary
In existing technologies, the production efficiency of Ganoderma lucidum spore powder is low, the cultivation cycle is long, and the contamination rate of the strain is high. It is impossible to break through the inefficient manual and workshop-style production mode. Moreover, the liquid fermentation strain technology is prone to causing large-scale pollution when applied in the field of edible and medicinal fungi.
A special solid-to-liquefaction microbial culture medium for Ganoderma lucidum cultivation on linden wood is used, which includes wheat starch, glucose, wheat cellulose, wheat bran, yeast extract, beef peptone, and potassium dihydrogen phosphate. The two-stage solid-to-liquefaction microbial culture method simplifies the process to include a mother culture and a special solid-to-liquefaction culture, shortening the culture cycle and avoiding the problems of expensive equipment and nutrient solution residue.
It significantly shortened the entire culture cycle of strain propagation, improved inoculation efficiency and spore yield, reduced production costs, decreased contamination rate and mutation risk, and achieved efficient and stable strain propagation and spore production.
Smart Images
Figure SMS_1 
Figure SMS_2 
Figure SMS_3
Abstract
Description
Technical Field
[0001] This invention relates to a method for culturing Ganoderma lucidum from linden wood and obtaining spore powder using a special solid-liquidation strain. Background Technology
[0002] Edible and medicinal fungi are one of the advantageous areas for the development of modern agriculture and the bio-industry, and China is one of the major producers and consumers of edible and medicinal fungi. Edible and medicinal fungi have a prominent position and excellent role in the transformation of modern biotechnology and the sustainable development of ecological agriculture, and play an irreplaceable role in social, economic and ecological development.
[0003] Currently, the edible and medicinal fungi industry system suffers from many factors that are incompatible with the development of new quality productive forces. The most representative of these is the lagging development of efficient breeding technology for the required strains, which has become a bottleneck restricting the industry's modernization. The conventional three-stage solid-state spawn breeding process commonly used in the industry is inefficient, has a long cultivation cycle, and a high rate of spawn contamination. It fails to break through the inefficient, manual, workshop-style production model, placing modern large-scale enterprises on the same competitive platform as small-scale spawn producers. This is the main root cause of frequent safety and quality incidents in edible and medicinal fungi production, and also the primary reason for the poor profitability of edible and medicinal fungi enterprises. In foreign countries such as Japan, liquid fermentation spawn technology is widely used in the industrialized production of edible and medicinal fungi. However, this technology has high requirements, large equipment investment, and is difficult to monitor online. Its application in the edible and medicinal fungi field is highly susceptible to large-scale contamination, leading to significant losses. Therefore, utilizing modern biotechnology and bioengineering techniques to achieve innovative and efficient spawn breeding is urgently needed.
[0004] Reishi mushroom is a type of large medicinal fungus. Modern medical research shows that reishi has pharmacological activities such as immunomodulation, anti-cancer, liver protection, sedation, and antioxidant effects. Red reishi mushroom. Ganoderma lucidum (Leyss. exFr.)Karst. and Purple Lingzhi Ganoderma sinense Zhao, Xu, and Zhang are all medicinal materials. In the field of Ganoderma lucidum spore powder production, spore powder obtained using the red Ganoderma lucidum linden wood production method is of high quality, high yield, and widely accepted by the market.
[0005] Cultivating Ganoderma lucidum on linden logs to obtain spore powder is the mainstream method in the Ganoderma lucidum spore powder industry, and it is also the product form with the best commercial quality and the highest market acceptance. Seed source and propagation techniques are key to linden log Ganoderma lucidum cultivation. Currently, the most commonly used method is a three-stage solid-state spawn propagation process (mother culture - bottled or bagged primary culture - bagged cultivation culture): The first stage is the mother culture, formulated with PDA modified medium (200g peeled potato, 20g glucose, 20g agar, 3g peptone, 1g potassium dihydrogen phosphate, 0.5g magnesium sulfate, 1000ml water), with a culture time of 9-15 days. One mother culture can be transferred to 3-5 bottles of primary culture. The second stage is the bottled or bagged primary culture, formulated mainly with sawdust (85% broadleaf tree sawdust, 14% wheat bran, 1% gypsum, moisture content 50-55%), with a culture time of 50-60 days. One bottle of primary culture can be transferred to 50-70 bottles of cultivation culture. The third stage is bagged spawn (1000ml~1200ml), with a formula mainly composed of sawdust medium (85% broadleaf tree sawdust, 14% wheat bran, 1% gypsum, moisture content 50~55%), and a culture time of 40~50 days. The entire culture cycle of the above conventional solid-state three-stage propagation process is 95~125 days.
[0006] Liquid fermentation cultures (also known as liquid submerged fermentation) have been used in a few cases in the cultivation of Ganoderma lucidum on linden wood. This process also involves a three-stage fermentation (mother culture - Erlenmeyer flask shaker fermentation liquid culture - fermentation tank submerged fermentation liquid culture): Stage 1 is the mother culture, with the same formula and preparation method as before, and a cultivation time of 9-15 days. One mother culture is used to inoculate 2-3 Erlenmeyer flask shaker fermentation liquid cultures (200ml each). Stage 2 is the Erlenmeyer flask shaker fermentation liquid culture (200ml of culture medium in a 500ml Erlenmeyer flask), with a formula mainly consisting of peeled fresh potatoes, glucose, yeast extract, peptone, etc., and a shaker cultivation time of 5-7 days. One liquid culture can be inoculated into 2000ml of the three-stage fermentation tank culture medium (10% inoculum). The third stage is a submerged fermentation liquid culture medium in a fermenter, with the same formula as the liquid primary culture medium in an Erlenmeyer flask and shaker. Fermentation requires a dedicated submerged fermenter or fermentation system, and the cultivation time is 4-5 days. 1 liter of liquid culture can be transferred to 30-50 linden wood bags. The entire cultivation cycle for the above liquid culture process is 18-27 days. Summary of the Invention
[0007] The technical problem to be solved by the present invention is to provide a method for obtaining spore powder from Ganoderma lucidum spores by culturing Ganoderma lucidum on linden wood using solid liquefied bacterial cultures.
[0008] To address the aforementioned technical problems, this invention provides a special culture medium for the solid-to-liquefaction of Ganoderma lucidum culturing on linden wood, which comprises the following components in parts by weight: Wheat starch 15-20 parts, glucose 1-2 parts, wheat cellulose 45-60 parts, wheat bran 15-25 parts, yeast extract 1-2 parts, beef peptone 1-2 parts, potassium dihydrogen phosphate 1-2 parts, water 100 parts.
[0009] The present invention also provides a method for preparing the above-mentioned culture medium, comprising the following steps: Mix wheat starch, bran, and wheat cellulose evenly to obtain ingredient I; Add glucose, yeast extract, beef peptone, and potassium dihydrogen phosphate to water and stir thoroughly (to dissolve glucose, yeast extract, beef peptone, and potassium dihydrogen phosphate in water) to obtain ingredient II; After thoroughly mixing ingredients I and II, a special culture medium for solid-liquid culture of Ganoderma lucidum on linden wood is obtained.
[0010] This invention also provides a method for obtaining spore powder from *Ganoderma lucidum* spores by culturing solid-liquid culture using the above-mentioned culture medium, comprising the following steps: 1) Solid-to-liquefied seed production; 2) Preparation of liquefied bacterial strains (bacterial solution); 3) Obtaining Ganoderma lucidum spore powder through inoculation, mycelium preparation, and cultivation.
[0011] An improvement to the method of obtaining spore powder from Ganoderma lucidum culturing on linden wood using solid liquefied bacterial cultures according to the present invention: The solid-liquid seed source preparation in step 1) is as follows: The solid-liquid culture medium for Ganoderma lucidum culture on linden wood was placed into a culture bottle, and after the cap was closed (after the microporous membrane vent cap was closed), it was sterilized (high temperature and high pressure sterilization at 121℃ and 0.11Mpa for 90 minutes) to obtain the sterilized culture medium; According to the inoculation amount of 6~6.5g (preferably 6g) of Ganoderma lucidum mother culture for every 200g of Ganoderma lucidum wood culture medium for solid-liquidation, the Ganoderma lucidum mother culture is inoculated into the sterilized culture medium under aseptic conditions, and cultured in the dark at 20~24℃ (preferably 22℃) until the mycelium fills the entire bottle (the culture time is generally 20~25 days), and the seed source (solid-liquidation special strain seed source) is obtained.
[0012] The preparation of the liquefied bacterial culture (bacterial solution) in step 2) is as follows: The seed source obtained in step 1) is first homogenized at high speed (homogenized and liquefied at a speed of 0.9~11,000 rpm for 0.8~1.2 minutes) under aseptic conditions to form a bacterial suspension. Then, it is diluted at a dilution ratio of 1:100 (which is a secondary dilution, i.e., 99 volumes of sterile water are added to 1 volume of bacterial suspension). The diluted product (pH naturally 5.5-6.5) is further liquefied in a dilution tank at a temperature of 20~22℃ and an aeration ratio of 1:0.4 (v / v / min) for 4~6 minutes (preferably 5 minutes) to obtain Ganoderma lucidum liquefied strain (Ganoderma lucidum liquefied strain liquid).
[0013] illustrate: 1. The mother culture of Ganoderma lucidum is obtained using conventional techniques; 2. To verify the purity and species characteristics of the liquefaction-specific microbial strain obtained in this invention, the liquefaction-specific microbial strains obtained in steps 1) and 2) can be subjected to the following tests: ① Purity testing, including mold testing and bacterial testing, using microscopic examination and routine bacterial testing standards; The types of molds tested are: Trichoderma and Penicillium; The bacteria tested were Bacillus subtilis.
[0014] ② Viability test, using the TTC method; ③ Sensory inspection: including antagonistic lines, absence of yellow water, and absence of discoloration; the culture bottle cap is intact and sealed, and the label is correct.
[0015] Step 3) involves inoculating the mycelium, preparing mycelium, and cultivating Ganoderma lucidum spore powder as follows: S1. Select suitable tree species as raw materials for mycelial linden culture; Cut branches with a diameter of 10cm-20cm into linden logs with a length of 25-30cm, controlling the moisture content to 38%-45%. Then, pack the linden logs into polyethylene tubes, placing 1-2cm of sawdust of the same tree species at both ends of the logs. Tie both ends of the polyethylene tubes with rope and immediately sterilize them at high temperature (sterilize at 98℃-100℃ under normal pressure for 16-24 hours). After the temperature of the linden logs is ≤30℃, under aseptic conditions, inoculate them with Ganoderma lucidum liquefied inoculum using the injection method, with an inoculation volume of 50-65ml / linden. Seal the injection port with medical tape to complete the preparation of the mycelium. S2. After inoculation, the mycelium is cultured at 15℃~22℃ in the dark for 60-70 days. When the surface of the mycelium segments turns light yellow, it is transferred to the beds in the cultivation field. The polyethylene tube bags outside the mycelium segments are removed, and the mycelium segments are arranged neatly with a spacing of 5cm~10cm and a row spacing of 20cm~25cm. The mycelium segments are covered with 0.5~1cm of soil. Finally, an arch frame is erected on the bed, with a height of 50cm~60cm, and covered with plastic film. During cultivation, maintain air humidity at 80%~95%, temperature at 20℃~27℃, good ventilation, and light intensity of 1000-2000 Lux, ensuring uniform illumination. After the Ganoderma primordia form, thin out each mycelium, retaining one Ganoderma fruiting body per mycelium. When the yellow growth ring on the outer edge of the Ganoderma cap disappears and spores begin to shoot out from under the cap, collect the spore powder by laying a film sleeve or covering the entire bed with cloth. After the Ganoderma spores have completely stopped shooting out, place the collected spore powder in a clean container and dry it at 45℃~50℃.
[0016] As a further improvement to the method of obtaining spore powder from *Ganoderma lucidum* culturing on linden wood using solid liquefied bacterial cultures according to the present invention, in step 3): Suitable tree species for linden culture are Fagaceae and Elaeocarpus.
[0017] The polyethylene tubular bag has a width of 30cm to 35cm, a thickness of 0.06mm to 0.08mm, and a length of 60cm to 80cm.
[0018] Therefore, this invention is a novel integrated method for cultivating Ganoderma lucidum on linden wood using a special solid-liquid culture medium, a method for preparing solid-liquid strains (seed source), a method for preparing liquefied strains (liquid), and a method for obtaining spores through cultivation.
[0019] The Ganoderma lucidum liquefaction culture obtained by this invention has abundant mycelial fragments and good dispersion (150 mycelial cells per field of view as detected by 400x microscopy). It is stable and has strong viability (TTC-dehydrogenase reduction method test: 0.2g of test sample + 2ml of 0.5% TTC-PBS (pH=8.0), stained in a constant temperature water bath at 40℃ for 2h, then 5ml of anhydrous ethanol was added for extraction at room temperature for 1h, absorbance of the extract OD465 value, note: 0.50-0.60 is acceptable). Satisfactory culture results can be obtained when the inoculum size is 50-65ml / linden.
[0020] Note: The inoculation volume of 50-65ml / linden refers to adding 50-65ml of bacterial solution (i.e., the Ganoderma lucidum liquefaction strain obtained in this invention) to each linden wood bag to be inoculated. After cultivation, the resulting product is Ganoderma lucidum spore powder from linden wood.
[0021] Compared with the conventional three-stage solid-liquid culture process, the Ganoderma lucidum solid-liquid culture method of the present invention has the following technical advantages: The strain propagation process of this invention is a two-stage process (mother culture - solid-to-liquefied special culture). The first step is the mother culture; the second step is the solid-to-liquefied special culture (200ml bottle of inoculum). The cultivation time is 20-25 days. When using, the solid-to-liquefied special culture only needs to be liquefied with sterile water (5 minutes) and can be directly used for inoculation of mycelium, without the need for a second cultivation process of mycelium. Therefore, the entire culture cycle of strain propagation only takes 30-35 days, which is significantly shortened by 65-90 days compared to the conventional three-stage solid culture process of 95-125 days. Each bottle of liquefied special culture (200g) is liquefied into 20 liters of bacterial solution, which can be used to inoculate 300 mycelium (50-65ml of bacterial solution per bag). Each mycelium contains 0.6g of solid culture, and the inoculation efficiency is more than 200 times that of conventional solid culture.
[0022] The biggest difference between the Ganoderma lucidum solid-liquidation special strain cultivation method of this invention and conventional liquid deep fermentation strains is that it adopts a two-stage seed source cultivation method, which is simple and does not require expensive stainless steel special fermentation equipment, saving more than 90% of investment. Because the seed source adopts independent small-package solid culture technology, the purity, viability and species characteristics of each bottle of seed source can be tested in real time before use, avoiding the inoculation risk caused by strain characteristics variation (the online testing of liquid strains propagated by conventional deep fermentation process is difficult before inoculation, and the species characteristics are difficult to control). Thirdly, the solid seed source of this invention only uses sterile water during liquefaction, without the need for a cultivation process. The bacterial liquid contains only pure mycelium and no nutrient solution residue, eliminating the problem of secondary bacterial contamination caused by nutrient solution issues in conventional liquid fermentation strains, improving the inoculation yield and the purity of the mycelium, and completely overcoming the drawbacks of liquid deep fermentation strains in inoculation applications.
[0023] The present invention has the following beneficial effects: The present invention provides a special solid-liquidation strain and liquefaction inoculation technology for Ganoderma lucidum, which simplifies the breeding process and improves propagation efficiency; reduces the amount of strain used, saving production costs; ensures strong mycelial vigor and controllable strain characteristics, reducing the risk of mutation; and produces uniform mycelial solution, meeting the trend of intelligent inoculation. Overall, this invention can reduce the inoculation cost by 60-70%, shortens the time for mycelial colony to fully develop after inoculation by about one-third compared to conventional solid-liquid strains, and increases spore yield to 180-210 grams per colony, which is 30-35% higher than conventional solid-liquid strains. The contamination rate is 0.2-0.4%, which is more than 100% lower than conventional solid-liquid strains, demonstrating significant cultivation advantages.
[0024] In summary, through integrated research on key technologies for the formation of Ganoderma lucidum strain quality and new technologies for large-scale and efficient propagation of Ganoderma lucidum spores on logs, this invention provides a special formula for solid-liquid Ganoderma lucidum strains, related cultivation techniques, and application technology for Ganoderma lucidum spore powder on logs. The solid-liquid strains produced using the method of this invention have a short cycle, vigorous mycelial growth, strong vitality, and low production cost. After liquefaction, they are used to cultivate Ganoderma lucidum spore powder on logs. After inoculation on logs, mycelium grows at multiple points, mycelial growth is rapid, and the yield is high. The inoculation efficiency is more than 100 times that of traditional solid strains. The yield and quality of spore powder are significantly improved. This invention is a new type of strain that is highly efficient, stable, and reliable, and a new model for the intensive cultivation of Ganoderma lucidum spore powder on logs. Detailed Implementation
[0025] Example 1: A special culture medium for solid-to-liquefied Ganoderma lucidum, which is composed of the following components in parts by weight: 15 parts wheat starch 1 serving of glucose 45 parts wheat cellulose 25 parts wheat bran 1 part yeast extract 1 serving of beef peptone 1 part potassium dihydrogen phosphate 100 portions of water.
[0026] The above-mentioned culture medium is prepared by performing the following steps in sequence: In a stainless steel container, wheat starch, wheat cellulose, and bran are mixed evenly under dry conditions to obtain ingredient I; Add glucose, yeast extract, beef peptone, and potassium dihydrogen phosphate to water and stir thoroughly (to fully dissolve glucose, yeast extract, beef peptone, and potassium dihydrogen phosphate in water) to obtain ingredient II; After thoroughly mixing ingredients I and II, a special culture medium for Ganoderma lucidum solid-liquidization is obtained.
[0027] Example 2: Using the culture medium (specific culture medium for solid-liquidation of Ganoderma lucidum) obtained in Example 1, a specific strain for solid-liquidation of Ganoderma lucidum was prepared by performing the following steps in sequence: 1) Production of special microbial strains (seed source) for solid liquefaction Immediately dispense 200g of the freshly prepared Ganoderma lucidum solid-liquidation special culture medium into 200ml special culture bottles, cover with a microporous membrane vent cap (using Shanghai Songtuo Industrial Co., Ltd. ZP14-200 vent culture bottle), and sterilize at 121℃ and 0.11Mpa for 90 minutes to obtain the sterilized culture medium; After the sterilized culture medium is cooled to below 25°C, 6 grams of Ganoderma lucidum mother culture (solid mother culture) is inoculated under aseptic conditions and cultured in the dark at 20°C for 25 days to obtain a solid-liquidation special strain (seed source); at this time, the mycelium has fully grown in the bottle.
[0028] The purity was tested and found to be 100%. Tests showed that no molds such as Trichoderma or Penicillium were detected. Tests showed that Bacillus subtilis and other bacteria were not detected. The viability data obtained by the TTC method was an OD485 value of 0.55; Sensory inspection results showed no primordia formation, the culture bottle cap was intact and sealed, and the label was correct.
[0029] 2) Preparation of liquefied bacterial strains (bacterial solution): The solid liquefaction-specific bacterial strain (inoculum) was first homogenized at high speed (homogenized and liquefied for 1 minute at a speed of 10,000 rpm) under aseptic conditions, and then diluted with sterile water at a dilution ratio of 1:100 (a secondary dilution). The resulting product (mycelial suspension, pH 6.5) was liquefied in a dilution tank at a liquefaction temperature of 20℃ and an aeration ratio of 1:0.4 (v / v / min) for 5 minutes to obtain the Ganoderma lucidum liquefaction bacterial strain (liquid).
[0030] Molds such as Trichoderma and Penicillium were not detected; bacteria such as Bacillus subtilis were not detected.
[0031] Example 3: Using the Ganoderma lucidum solid-liquidation special inoculum liquid obtained in Example 2, the following steps were performed sequentially to prepare mycelial substrate and obtain spore powder through cultivation: S1. Select oak trees (Quercus variabilis) of the Fagaceae family, with a diameter of 10cm~20cm, cut into 25cm lengths per linden, and control their moisture content to 38%~45%. Pack the linden wood into polyethylene tubular bags (30cm~35cm wide, 0.06mm~0.08mm thick, and 60cm~80cm long). Place 1~2cm of oak wood chips at both ends of the linden wood. Tie both ends of the bag with rope and immediately sterilize at 98℃~100℃ under normal pressure for 24 hours. After the temperature of the linden wood is ≤26℃, inoculate with liquefied inoculum (bacterial solution) under aseptic conditions using the injection method. The inoculation volume is 50~65ml / linden. Seal the inoculation (injection) port with medical adhesive tape to complete the preparation of mycelial linden.
[0032] S2. After inoculation, the mycelium segments are cultured at 15℃~22℃ in the dark for 60-70 days. When a light yellow mycelial skin appears on the surface of the segments, they are transferred to the beds in the cultivation area. The polyethylene bags outside the mycelium segments are removed, and the segments are arranged neatly with a spacing of 5cm~10cm between segments and a row spacing of 20cm~25cm. The segments are covered with 0.5cm~1cm of soil. Finally, an arched frame is erected on the bed, with a height of 50cm~60cm, and covered with plastic film.
[0033] During cultivation, maintain air humidity at 80%~95%, temperature at 20℃~27℃, good ventilation, and uniform light at 1000~2000 Lux. After the Ganoderma primordia have formed, thin out each mycelium, leaving one Ganoderma per mycelium. When the yellow growth ring on the cap disappears and a small number of spores are released from under the cap, collect the spore powder by covering the entire bed with cloth. After the Ganoderma stops releasing spores, place the spore powder in a clean container and dry it at 45℃~50℃. This completes the spore powder acquisition process.
[0034] The results of comparing the Ganoderma lucidum solid-liquidation strain of the present invention with those of conventional solid and liquid fermentation strains obtained by inoculating with the same inoculum and harvesting Ganoderma lucidum spore powder are as follows: Table 1. Comparison of the inoculation effects of Ganoderma lucidum liquefaction strains, solid strains, and liquid fermentation strains on inoculum.
[0035] Comparing the data in Table 1 above, it can be seen that using the solid-liquid Ganoderma lucidum strain of the present invention to inoculate Ganoderma lucidum linden wood bags to obtain spore powder is far superior to the results obtained by conventional techniques.
[0036] Comparative Example 1-1: The formulation of the Ganoderma lucidum solid-to-liquefaction special culture medium in Example 1 was modified as follows: The use of 45 parts wheat cellulose was omitted, and the wheat starch was increased from 15 parts to 30 parts accordingly; the rest was the same as in Example 1.
[0037] Comparative Examples 1-2: The formulation of the Ganoderma lucidum solid-to-liquefaction special culture medium in Example 1 was modified as follows: Replace “45 parts wheat cellulose” with “45 parts lignin”, and the rest is the same as in Example 1.
[0038] Comparative Examples 1-3: The formulation of the Ganoderma lucidum solid-to-liquefaction special culture medium in Example 1 was modified as follows: Replace “45 parts wheat cellulose” with “45 parts fine wood chips”, and the rest is the same as in Example 1.
[0039] The culture media obtained in Comparative Examples 1-1 to 1-3 were used for the Ganoderma lucidum solid-liquidation special strain culture method described in Example 2. The resulting Ganoderma lucidum solid-liquidation special strain culture solution was then used in Example 3 to obtain spore powder. The time required for the mycelium to fully colonize the bottle is shown in Table 2. The comparison of the results is shown in Table 2 below: Table 2
[0040] Comparing the data in Table 2 above, it can be seen that the inoculation of the solid liquefied Ganoderma lucidum strain of the present invention with linden fungi for cultivation yields spore powder significantly better than that of Comparative Examples 1-1, 1-2 and 1-3.
[0041] Comparative Example 2-1: The following changes are made to "2), Preparation of liquefied bacterial strain (bacterial solution):" in Example 2: The dilution ratio was changed from "1:100" to "1:1"; the rest was the same as in Example 2.
[0042] The result was that the bacterial strain could not be liquefied in the subsequent step 2) and became a viscous substance. The liquefaction of the bacterial strain failed.
[0043] Comparative Example 2-2: The following changes are made to "2) Preparation of liquefied bacterial strain (bacterial solution):" in Example 2: Change the dilution ratio from "1:100" to "1:200"; the rest is the same as in Example 2.
[0044] Comparative Examples 2-3: The following changes were made to "2) Preparation of liquefied bacterial strain (bacterial solution):" in Example 2: The ventilation ratio was changed from "1:0.4" to "1:0.2"; the rest is the same as in Example 2.
[0045] Comparative Examples 2-4: The following changes were made to "2) Preparation of liquefied bacterial strain (bacterial solution):" in Example 2: The ventilation ratio was changed from "1:0.4" to "1:0.8"; the rest is the same as in Example 2.
[0046] Comparative Example 3-1: The following changes are made to "1) Preparation of special strains (seed source) for solid liquefaction" in Example 2: The culture temperature was changed from "20℃" to "22℃"; the rest is the same as in Example 2.
[0047] Comparative Example 3-2: The following changes are made to "1) Preparation of special strains (seed source) for solid liquefaction" in Example 2: The culture temperature was changed from "20℃" to "18℃"; the rest is the same as in Example 2.
[0048] Using the Ganoderma lucidum solid-liquefaction special inoculum liquids obtained in Comparative Examples 2-2~2-4 and Comparative Examples 3-1~3-2, the mycelium was inoculated into linden trees and cultivated according to the method described in Example 3. Finally, spore powder was obtained, and the results are compared in Table 3 below: Table 3
[0049] Comparing the data in Table 3 above, it can be seen that the inoculation of the solid-liquid Ganoderma lucidum strain of the present invention with linden fungi and cultivation results in significantly better spore powder production than comparative examples 2-1~2-4 and 3-1~3-2.
[0050] Comparative Example 4-1: The amount of bacterial solution used to inoculate the linden fruiting body with the Ganoderma lucidum solid-liquefaction special strain in Example 3 was modified from 50-65 ml / linden as follows: The inoculation volume of the bacterial solution was changed from "50~65ml / linden" to "25~35ml / linden"; the rest was the same as in Example 3.
[0051] Comparative Example 4-2: The inoculation volume of the bacterial solution was changed from "50~65ml / linden" to "100~130ml / linden"; the rest was the same as in Example 3.
[0052] The results obtained are compared with those of Example 3 in Table 4 below: Table 4
[0053] As can be seen from the data comparison in Table 4 above, the present invention, which uses conventional methods to inoculate linden fungi for cultivation and finally obtains spore powder, has a significantly better overall effect than comparative examples 4-1 to 4-2.
[0054] Finally, it should be noted that the above examples are merely some specific embodiments of the present invention. Obviously, the present invention is not limited to the above embodiments and many variations are possible. All variations that can be directly derived or conceived by those skilled in the art from the disclosure of the present invention should be considered within the scope of protection of the present invention.
Claims
1. A special culture medium for solid-to-liquefaction microbial strains for culturing Ganoderma lucidum on linden wood, characterized in that... It consists of the following components in parts by weight: Wheat starch 15-20 parts, glucose 1-2 parts, wheat cellulose 45-60 parts, wheat bran 15-25 parts, yeast extract 1-2 parts, beef peptone 1-2 parts, potassium dihydrogen phosphate 1-2 parts, water 100 parts.
2. The method for preparing the culture medium according to claim 1, characterized in that... Includes the following steps: Mix wheat starch, bran, and wheat cellulose evenly to obtain ingredient I; Add glucose, yeast extract, beef peptone, and potassium dihydrogen phosphate to water and stir thoroughly to obtain ingredient II; After thoroughly mixing ingredients I and II, a special culture medium for solid-liquid culture of Ganoderma lucidum on linden wood is obtained.
3. A method for obtaining spore powder from *Ganoderma lucidum* spores by culturing solid-liquid cultured with the culture medium as described in claim 1, characterized in that: Perform the following steps in sequence: 1) Solid-to-liquefied seed production; 2) Preparation of liquefied bacterial strains; 3) Obtaining Ganoderma lucidum spore powder through inoculation, mycelium preparation, and cultivation.
4. The method for obtaining spore powder from *Ganoderma lucidum* culturing on linden wood using solid-liquid culture according to claim 3, characterized in that... The solid-liquid seed source preparation in step 1) is as follows: The solid-liquid culture medium for Ganoderma lucidum culture on linden wood is placed into a culture bottle, the cap is closed, and then sterilized to obtain the sterilized culture medium. According to the inoculation amount of 6~6.5g of Ganoderma lucidum mother culture for every 200g of Ganoderma lucidum wood culture medium, the Ganoderma lucidum mother culture was inoculated into the sterilized culture medium under aseptic conditions, and cultured in the dark at 20~24℃ until the mycelium filled the whole bottle to obtain the seed source.
5. The method for obtaining spore powder from *Ganoderma lucidum* culturing on linden wood using solid-liquid culture according to claim 4, characterized in that... The preparation of the liquefied bacterial strain in step 2) is as follows: The seed source is first homogenized into a bacterial suspension under sterile conditions using high speed, and then diluted at a dilution ratio of 1:
100. The diluted product is then liquefied in a dilution tank at a temperature of 20~22℃ and an aeration ratio of 1:0.4 (v / v) for 4~6 minutes to obtain Ganoderma lucidum liquefaction strain.
6. The method for obtaining spore powder from *Ganoderma lucidum* culturing on linden wood using solid-liquid culture according to claim 5, characterized in that... The steps in step 3) of inoculating the bacterial solution, preparing the mycelium, and cultivating Ganoderma lucidum spore powder are as follows: S1. Select suitable tree species as raw materials for mycelial linden culture; Cut branches with a diameter of 10cm-20cm into linden logs with a length of 25-30cm, control the moisture content to 38%-45%, then put the linden logs into polyethylene tube bags, place 1-2cm of sawdust of the same tree species at both ends of the linden logs, tie both ends of the polyethylene tube bags with rope, and immediately sterilize at high temperature; after the temperature of the linden logs is ≤30℃, under aseptic conditions, inoculate with Ganoderma lucidum liquefied inoculum using the injection method, with an inoculation volume of 50-65ml / linden, and seal the inoculation injection port with medical tape to complete the preparation of mycelial lindens; S2. After inoculation, the mycelium is cultured at 15℃~22℃ in the dark for 60-70 days. When the surface of the mycelium segments turns light yellow, it is transferred to the beds in the cultivation field. The polyethylene tube bags outside the mycelium segments are removed, and the mycelium segments are arranged neatly with a spacing of 5cm~10cm and a row spacing of 20cm~25cm. The mycelium segments are covered with 0.5~1cm of soil. Finally, an arch frame is erected on the bed, with a height of 50cm~60cm, and covered with plastic film. During cultivation, maintain air humidity at 80%~95%, temperature at 20℃~27℃, good ventilation, and light intensity of 1000~2000 Lux, ensuring uniform illumination. After the Ganoderma primordia form, thin out each mycelium, retaining one Ganoderma fruiting body per mycelium. When the yellow growth ring on the outer edge of the Ganoderma cap disappears and spores begin to shoot out from under the cap, collect the spore powder by laying a film sleeve or covering the entire bed with cloth. After the Ganoderma spores have completely stopped shooting out, place the collected spore powder in a clean container and dry it at 45℃~50℃.
7. The method for obtaining spore powder from *Ganoderma lucidum* culturing on linden wood using solid-liquid culture according to claim 6, characterized in that... In step 3): Suitable tree species for linden culture are Fagaceae and Elaeocarpus.
8. The method for obtaining spore powder from *Ganoderma lucidum* spores by culturing *Ganoderma lucidum* on linden wood using solid-liquid culture according to claim 7, characterized in that... In step 3): Polyethylene tubular bags have a width of 30cm to 35cm, a thickness of 0.06mm to 0.08mm, and a length of 60cm to 80cm.