Method for inducing rapid formation of boletus sinensis primordium

Abstract

The invention discloses a method for inducing rapid formation of Chinese saprophytic boletus primordium, and belongs to the technical field of edible fungus cultivation. The method comprises the following steps: activating a boletus sinensis strain JSJ-Bx1, inoculating the boletus sinensis strain JSJ-Bx1 into a culture medium, and carrying out dark culture; the preservation number of the boletus sinensis strain JSJ-Bx1 is CGMCC (China General Microbiological Culture Collection Center) No.42007, and the boletus sinensis strain JSJ-Bx1 is preserved in the China General Microbiological Culture Collection Center; performing liquid fermentation culture on the hyphae which are activated and are almost overgrown on the flat plate to obtain fermentation liquid; inoculating a cultivation material with the fermentation liquid, performing dark culture until the bottle / bag is full of hyphae, covering the bottle / bag full of the hyphae with soil, covering the surface with non-woven fabric as a protective layer, and performing hypha culture and light culture for 3-5 days to obtain a soil-covered fruiting bottle / bag; performing light culture on the soil-covered fruiting bottle / bag in a fruiting room, and spraying a monopotassium phosphate solution during culture to promote mycelial growth and induce primordium formation. The method can effectively promote rapid growth of Chinese saprophytic boletus hyphae in an overburden layer, primordia are rapidly formed on the surface of the overburden layer, and harvesting can be performed 13-15 days after overburden.

Description

Technical Field

[0001] This invention belongs to the field of edible fungi cultivation technology, specifically relating to a method for rapidly inducing the formation of primordia of Boletus sinensis. Background Technology

[0002] Chinese Boletus saprophyticus Buchwaldoboletus xylophilus (Petch) Both&B. Ortiz, belonging to the family Boletaceae, subfamily Chalciporoideae, genus *Boletus*. Buchwaldoboletus It is a typical tropical and subtropical rare bolete, a non-ectomycorrhizal fungus with saprophytic ability, and is edible.

[0003] As the second newly discovered bolete species capable of artificial cultivation, *Boletus saprophyticus* possesses significant scientific and commercial value. However, basic biological research on *Boletus saprophyticus* is currently weak. There are few reports on its biological characteristics, domestication, cultivation, and molecular biology, and the current artificial cultivation technology system is still incomplete. For example, current authorized or published patents report that primordia only form 10–15 days after the surface of the casing layer develops a large number of golden-yellow mycelia; this excessively long primordia formation time prolongs the cultivation cycle. Furthermore, *Boletus saprophyticus* is a typical thermophilic fungus, making it highly susceptible to contamination in high-temperature and high-humidity cultivation environments.

[0004] References are available at: [1] Yunnan Tropical Crops Research Institute. An artificial cultivation method for Boletus sinensis: CN113348963A[P]. [2]Nanu S, Kumar TKA, 2022. Buchwaldoboletus xylophilus , a boletoidfungus new to India. Mycology, 7(1): 95-98. [3]Xie HJ, Zhang CX, He MX, Liang ZQ, Deng XH, Zeng NK, 2021. Buchwaldoboletus xylophilus and Phlebopus portentosus, two non-ectomycorrhizal boletes from tropical China. Phytotaxa, 520(2): 137-154. Summary of the Invention

[0005] The purpose of this invention is to address the problems of late primordia formation and high contamination rate of *Boletus spp.* by providing a method to rapidly induce primordia formation, effectively promoting mycelial growth and rapidly inducing primordia formation, significantly shortening the cultivation cycle and reducing production costs. Simultaneously, the rapid mycelial growth and primordia formation effectively resist the growth of other microorganisms, reducing the contamination rate of fruiting bottles / bags and increasing fruiting body yield and economic benefits.

[0006] The technical solution adopted in this invention is as follows: A species of Boletus sinensis that induces saprophytic growth ( Buchwaldoboletus xylophilus A method for rapid primordium formation includes the following steps: The first step involved activating the *Boletus saprophyticus* strain JSJ-Bx1 and inoculating it into B1 medium for dark incubation for 5 days at 30–35°C. The *Boletus saprophyticus* strain JSJ-Bx1 has the accession number CGMCC No. 42007 and is deposited at the China General Microbiological Culture Collection Center. The B1 medium consisted of the following components: 10–30 g glucose, 2–6 g yeast extract, 1–3 g ferrous sulfate, 0.1–0.3 g 1 / 2 MS medium, 18–20 g agar, and 1 L water.

[0007] The second step is to carry out liquid fermentation culture on the activated mycelium that is about to cover the plate. The culture temperature is 28–30℃ and the culture is carried out in the dark for 7 days to obtain the fermentation liquid. The third step is to inoculate the fermentation liquid into the cultivation medium and incubate it in the dark for 30 days until the mycelium fills the bottle / bag. The incubation temperature is 28–30℃. The fourth step is to cover the bottles / bags filled with mycelium with soil, and then cover the surface with non-woven fabric as a protective layer for mycelium cultivation in the soil layer. After 3-5 days of cultivation, the bottles / bags with soil covering and fruiting are obtained. The fifth step is to place the mushroom bottles / bags covered with soil into a mushroom house at 26–28℃ for light cultivation. During cultivation, spray with potassium dihydrogen phosphate solution to promote mycelial growth and induce primordia formation.

[0008] Preferably, in the fourth step above, the nonwoven fabric has a density of 20 g / m². 2 The non-woven fabric is covered with soil with a moisture content of 50–60% and a thickness of 1–3 cm.

[0009] Preferably, the mass concentration of the potassium dihydrogen phosphate solution used in the fifth step above is 0.05–0.3%.

[0010] Preferably, the moisture content of the covering material is 55%, the covering thickness is 1.5 cm, and the mass concentration of the potassium dihydrogen phosphate solution is 0.2%.

[0011] This invention effectively promotes the rapid growth of *Boletus chinensis* mycelium in the casing layer, achieving full coverage in just 2-3 days and rapid formation of primordia on the surface. Primordia formation is induced in just 3-5 days, and harvesting can begin 13-15 days after casing. Compared to the latest reports that require 10-15 days for primordia formation after casing, this significantly shortens the primordia formation time, reduces the overall cultivation cycle, and lowers production costs. Furthermore, the rapid mycelial coverage and primordia formation effectively resist the growth of other microorganisms, reduces contamination rates in fruiting bottles / bags, increases fruiting body yield, improves economic efficiency, and is more suitable for industrialized production. Attached Figure Description

[0012] Figure 1 The growth of *Boletus saprophyticus* JSJ-Bx1 after 5 days of culture under different primordium induction treatments; Figure 2 The growth of *Boletus saprophyticus* JSJ-Bx1 after 7 days of culture under different primordium induction treatments; Figure 3 The growth of *Boletus saprophyticus* JSJ-Bx1 after 10 days of culture under different primordium induction treatments; Figure 4 The growth of *Boletus chinensis* JSJ-Bx1 after 14 days of culture under different primordium induction treatments; Figure 5 The growth of *Boletus chinensis* JSJ-Bx1 after 4 days of induction culture by spraying with potassium dihydrogen phosphate solution of different concentrations; Figure 6 The growth of *Boletus chinensis* JSJ-Bx1 induced by spraying with potassium dihydrogen phosphate solution of different concentrations for 6 days; Figure 7 This describes the primordium induction and fruiting body growth in Example 1. Figure 8 This describes the primordium induction and fruiting body growth in Example 2. Figure 9 This shows the primordium induction and fruiting body growth in Example 3. Detailed Implementation

[0013] To make the objectives, technical solutions, and advantages of this invention clearer, the following detailed description of the invention is provided in conjunction with specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of protection of this invention.

[0014] (1) Preparation of fruiting bottles / bags for *Boletus chinensis* After activation, the *Boletus chinensis* strain JSJ-Bx1 was inoculated into B1 medium and cultured. The activated mycelium was cultured at 30–35℃ for 5 days.

[0015] The strain JSJ-Bx1 of the mother culture of *Boletus spp.* is deposited at the China General Microbiological Culture Collection Center (CGMCC No. 42007) under the classification name *Boletus spp.* Buchwaldoboletus xylophilus The deposit date is May 15, 2025.

[0016] The parent strain of the *Boletus chinensis* strain was collected on June 5, 2024, from wild fruiting bodies in the forest under the trees in Jinghong City, Xishuangbanna Dai Autonomous Prefecture. Pure culture mycelia were obtained through tissue isolation, purification, and identification.

[0017] The B1 culture medium has the following mass ratio: glucose 10–30 g, yeast extract 2–6 g, ferrous sulfate 1–3 g, 1 / 2 MS medium 0.1–0.3 g, agar 18–20 g, and water 1 L. A preferred ratio is: glucose 20 g, yeast extract 2 g, ferrous sulfate 1.5 g, 1 / 2 MS medium 0.3 g, agar 20 g, and water 1 L.

[0018] After activation, the mycelia that have almost fully colonized the plate are taken in half-plates, chopped, and then subjected to liquid fermentation at 28–30°C for 7 days in the dark. The liquid fermentation medium has the following mass ratio: glucose 10–30 g, yeast extract 2–6 g, ferrous sulfate 1–3 g, 1 / 2 MS medium 0.1–0.3 g, and water 1 L. An optimal ratio is: glucose 20 g, yeast extract 2 g, ferrous sulfate 1.5 g, 1 / 2 MS medium 0.3 g, and water 1 L.

[0019] The fermentation liquid is inoculated into the cultivation substrate. After inoculation, the mycelium culture temperature of the fruiting bottles / bags is 28–30℃, and cultured for 30 days until the mycelium fills the bottle / bag. The cultivation substrate ratio is: sawdust 20–50%, wheat grains 20–40%, wheat bran 5–30%, cottonseed hulls 10–30%, corn flour 5–10%, gypsum powder 0.05–0.1%, and potassium dihydrogen phosphate 0.05–0.1%. A preferred cultivation substrate formula is: sawdust 40%, wheat grains 20%, wheat bran 10%, cottonseed hulls 20%, corn flour 10%, gypsum powder 0.1%, and potassium dihydrogen phosphate 0.1%.

[0020] (2) Screening of the thickness of the soil covering layer for *Boletus chinensis* After mixing equal volumes of red soil and peat soil, add water and stir, controlling the moisture content to 55%, to obtain the covering material.

[0021] Three different treatments were set up: a 3 cm soil covering layer and spraying with water; a 3 cm soil covering layer and spraying with a 0.1% potassium dihydrogen phosphate solution; and a 1.5 cm soil covering layer and spraying with water. After soil covering, mushroom cultivation management was carried out uniformly, with induction culture lasting 5, 7, 10, and 14 days, respectively. Results are shown below. Figures 1-4 .

[0022] Figure 1 This image shows a comparison of the growth of *Boletus sinensis* JSJ-Bx1 after 5 days of induction culture on different primordia. The left image shows a soil covering thickness of 3 cm and spraying with water; the middle image shows a soil covering thickness of 3 cm and spraying with 0.1% potassium dihydrogen phosphate solution; and the right image shows a soil covering thickness of 1.5 cm and spraying with water.

[0023] Figure 2 This image shows a comparison of the growth of *Boletus sinensis* JSJ-Bx1 after 7 days of induction culture on different primordia. The left image shows a soil covering thickness of 3 cm and spraying with water; the middle image shows a soil covering thickness of 3 cm and spraying with 0.1% potassium dihydrogen phosphate solution; and the right image shows a soil covering thickness of 1.5 cm and spraying with water.

[0024] Figure 3 This image shows a comparison of the growth of *Boletus chinensis* JSJ-Bx1 after 10 days of induction culture on different primordia. The left image shows a soil covering thickness of 3 cm with water spraying; the middle image shows a soil covering thickness of 3 cm with 0.1% potassium dihydrogen phosphate solution spraying; and the right image shows a soil covering thickness of 1.5 cm with water spraying.

[0025] Figure 4 This image shows a comparison of the growth of *Boletus chinensis* JSJ-Bx1 after 14 days of induction culture on different primordia. The left image shows a soil covering thickness of 3 cm with water spraying; the middle image shows a soil covering thickness of 3 cm with 0.1% potassium dihydrogen phosphate solution spraying; and the right image shows a soil covering thickness of 1.5 cm with water spraying.

[0026] As shown in the figure, the thickness of the casing layer significantly affected mycelial growth and primordium formation. In the treatment with a casing layer thickness of 1.5 cm and water spraying, the mycelium quickly (within 2–3 days) covered the casing layer, and primordia formed on days 4–5 after casing, with primordia ranging in size from 0.5 to 1.5 cm. By day 7 after casing, the primordia had completely covered the casing layer, with a large number of primordia (10–20) and a size of 1–3 cm. By day 10 after casing, the primordia had clearly differentiated into fruiting bodies, with a relatively large number of 5–10 fruiting bodies, a size of 3–5 cm. By day 14 after casing, the fruiting bodies reached the harvest standard, with cap diameter of 6–8 cm, stipe length of 8–12 cm, stipe diameter of 2–4 cm, yield per bottle / bag of 100–120 g, and a biological conversion efficiency of 31.75–38.10%.

[0027] When the casing layer is 3 cm thick and water is sprayed, the mycelium takes a long time to fully grow into the casing layer, requiring 7–10 days. On the 8th–10th day after casing, some fruiting bottles / bags show primordia formation, with a small number of 2–3 primordia, each 1–2 cm in size. In addition, most fruiting bottles / bags have green mold contamination on the surface of the casing layer, making it difficult for the fruiting bodies to develop and grow. At harvest, the cap diameter is 3–5 cm, the stipe length is 4–5 cm, the stipe diameter is 2–3 cm, the yield per bottle / bag is 50–60 g, and the biological conversion efficiency is 15.87–19.05%.

[0028] Therefore, if only water is sprayed and the casing layer is 1.5 cm thick, it is beneficial for the mycelium of *Boletus sinensis* to quickly fill the casing layer and rapidly form primordia to enter the fruiting body development stage. At the same time, this method results in dense mycelium in the casing layer without water leakage, rapid mycelial filling of the casing layer, strong resistance to contamination on the casing layer surface, and no pollution. The primordia differentiation time is short, only 3-5 days, which significantly shortens the cultivation cycle. Fruiting bodies can be harvested 13-15 days after casing. The yield of fruiting bodies per bottle / bag is high, and the biological transformation efficiency is high.

[0029] (3) Screening of the concentration of fruiting water from *Boletus edulis* (3.1) Mix equal volumes of red soil and peat moss, then add water and stir to achieve a casing material moisture content of 55% and a casing thickness of 3 cm. After casing, manage the mushroom growth uniformly. Prepare a 0.10% potassium dihydrogen phosphate solution and spray 5 mL of the solution per bottle / bag daily for primordia induction. Use 5 mL of water per bottle / bag daily as a control. Results are shown in […]. Figures 1-4 .

[0030] As shown in the figure, daily spraying with 0.1% potassium dihydrogen phosphate solution significantly promoted mycelial growth and induced primordium differentiation of *Boletus sinensis*. The treatment with potassium dihydrogen phosphate solution resulted in rapid mycelial growth, almost completely covering the casing layer by day 5 after casing, and primordia forming by day 7. The number of primordia was relatively small, ranging from 3 to 5, with a size of 1–3 cm. By day 10 after casing, the primordia had clearly differentiated into fruiting bodies, with a number of 2 to 5, and a size of 2–4 cm. The fruiting bodies were ready for harvest by day 13 after casing, with cap diameter of 5–7 cm, stipe length of 8–10 cm, and stipe diameter of 2–3 cm. The yield per bottle / bag was 100–110 g, and the biological conversion efficiency was 31.75–34.92%.

[0031] The control group, which was sprayed with water after covering with 3 cm of soil, experienced slow mycelial growth, requiring 7–10 days to fully cover the soil layer. Primordia formed on some fruiting bottles / bags 8–10 days after covering, but the number of primordia was small, only 2–3, with a size of 1–2 cm. Furthermore, most fruiting bottles / bags had green mold contamination on the surface of the soil layer. The fruiting bodies were difficult to develop and grow, with cap diameter of 3–5 cm, stipe length of 4–5 cm, and stipe diameter of 2–3 cm at harvest. The yield per bottle / bag was 50–60 g, and the biological conversion efficiency was 15.87–19.05%.

[0032] The above results indicate that spraying potassium dihydrogen phosphate solution after covering Boletus sinensis with soil can significantly promote mycelial growth in the soil layer and accelerate primordium induction.

[0033] (3.2) To further explore the optimal concentration of potassium dihydrogen phosphate solution sprayed during primordium induction after casing of *Boletus sinensis*, a single-factor experiment was conducted. Equal volumes of red soil and peat moss were mixed, and water was added to achieve a casing material moisture content of 55%. A casing thickness of 1.5 cm was selected, and mushroom cultivation management was implemented uniformly after casing. Potassium dihydrogen phosphate solutions with mass concentrations of 0% (i.e., water), 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, and 0.30% were prepared. 5 mL of each bottle / bag of potassium dihydrogen phosphate solution at different concentrations was sprayed daily for primordium induction. The results are shown in […]. Figure 5 , Figure 6 .

[0034] Figure 5 The growth of *Boletus sinensis* JSJ-Bx1 after 4 days of induction culture with different concentrations of potassium dihydrogen phosphate solution is shown in the images. The seven groups of samples from left to right are the results of spraying with potassium dihydrogen phosphate solution at concentrations of 0%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, and 0.30%, respectively. Figure 6The growth of *Boletus sinensis* JSJ-Bx1 after 6 days of induction culture with different concentrations of potassium dihydrogen phosphate solution is shown in the images. The seven groups of samples from left to right are the results of spraying with potassium dihydrogen phosphate solution at concentrations of 0%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, and 0.30%, respectively.

[0035] from Figure 5 It can be seen that on the 4th day after covering with soil, the surface of the soil layer treated with 0.15% and 0.20% potassium dihydrogen phosphate solution was almost covered with mycelium, and primordia were differentiating. Among them, the surface of the soil layer treated with 0.15% potassium dihydrogen phosphate solution was almost covered with mycelium, with 1–3 primordia and a size of 0.5–1.5 cm. The surface of the soil layer treated with 0.20% potassium dihydrogen phosphate solution was covered with mycelium, the primordia differentiated earlier, and there were more primordia, with 2–3 primordia and a size of 1–2 cm. Mycelia were clearly visible in the casing layer after spraying with 0.05%, 0.10%, and 0.25% potassium dihydrogen phosphate solution, but no primordia were differentiated. However, no mycelia were observed on the surface of the casing layer after spraying with 0.00% potassium dihydrogen phosphate solution or 0.30% potassium dihydrogen phosphate solution. This may be because the concentration of potassium dihydrogen phosphate solution was too high, which inhibited mycelial growth.

[0036] from Figure 6 It can be seen that on the 6th day after covering with soil, the surface of the soil layer treated with 0.00% potassium dihydrogen phosphate solution began to differentiate into primordia, with a small number of primordia and a size of 0.5–1.5 cm; the primordia of the treatment treated with 0.20% potassium dihydrogen phosphate solution gradually developed into fruiting bodies, with larger primordia and a size of 2–3 cm; no mycelia were observed on the surface of the soil layer of the treatment treated with 0.30% potassium dihydrogen phosphate solution, but primordia formed in a few fruiting bottles, with a size of 0.5–1 cm.

[0037] The above results indicate that the thickness of the casing layer significantly affects the complete coverage of the casing layer after the *Boletus spp.* JSJ-Bx1 strain is covered. In production trials, a casing layer thickness of 1.5 cm is considered optimal, as it significantly promotes rapid mycelial growth and primordia formation, reduces contamination, and increases fruiting body yield. Furthermore, spraying with potassium dihydrogen phosphate solution also significantly promotes mycelial growth in the casing layer and accelerates primordia induction, leading to rapid fruiting body development. The optimal concentration of potassium dihydrogen phosphate solution is 0.20%, which effectively promotes rapid mycelial growth and primordia differentiation from vegetative to reproductive growth, shortening the cultivation cycle and increasing fruiting body yield. Additionally, a casing layer thickness of 3 cm, combined with simultaneous spraying of an appropriate concentration of potassium dihydrogen phosphate solution, results in a shorter primordia induction time and rapid fruiting body development, which can be further applied to production cultivation to shorten the cultivation cycle, reduce production costs, and improve economic benefits. Example 1

[0038] The *Boletus chinensis* strain JSJ-Bx1 was activated and inoculated into B1 medium for culture. The activated mycelium was cultured at 30–35℃ for 5 days. The B1 medium composition was: 20 g glucose, 2 g yeast extract, 1.5 g ferrous sulfate, 0.3 g 1 / 2 MS medium, 20 g agar, and 1 L water. The activated mycelium, having fully colonized the plates, was then chopped and subjected to liquid fermentation at 28–30℃ for 7 days in the dark. The liquid fermentation medium consisted of: 20 g glucose, 2 g yeast extract, 1.5 g ferrous sulfate, 0.3 g 1 / 2 MS medium, and 1 L water. The fermentation liquid was inoculated into the substrate. After inoculation, the mycelium in the fruiting bottles / bags was cultured at 28–30℃ for 30 days until the mycelium fully colonized the bottles / bags. The cultivation substrate formula was: 40% sawdust, 20% wheat grains, 10% wheat bran, 20% cottonseed hulls, 10% corn flour, 0.1% gypsum powder, and 0.1% potassium dihydrogen phosphate. Equal volumes of red soil and peat moss were mixed, and water was added to achieve a moisture content of 50%. The casing layer was set at a thickness of 3 cm. After casing, water was sprayed daily for mushroom cultivation management, and the results were recorded. (See attached table for details.) Figure 7 .

[0039] Figure 7 The left-hand diagram shows the primordium induction and fruiting body growth in this embodiment, while the right-hand diagram shows the fruiting body at harvest time.

[0040] Process records show that it takes 8–10 days for the mycelium of *Boletus sinensis* to fully colonize the casing layer, and 11–15 days for primordia formation. The number of primordia is low, ranging from 1 to 3, and their size is 1–2 cm. The casing layer surface is contaminated with other microorganisms. At harvest, the cap diameter is 3–5 cm, the stipe length is 4–5 cm, and the stipe diameter is 2–3 cm. The yield per bottle / bag is 40–60 g, with a biological conversion efficiency of 12.70–19.05%. This method results in a long time for the mycelium to fully colonize the casing layer, a low number of primordia, and most primordia becoming contaminated with *Phyllostachys edulis* during development, leading to low economic efficiency. Example 2

[0041] The *Boletus chinensis* strain JSJ-Bx1 was activated and inoculated into B1 medium for 5 days at 30–35℃. The B1 medium composition was: 20 g glucose, 2 g yeast extract, 1.5 g ferrous sulfate, 0.3 g 1 / 2 MS medium, 20 g agar, and 1 L water. Half a dish of activated mycelium that had fully colonized the plates was then chopped and subjected to liquid fermentation at 28–30℃ in the dark for 7 days. The liquid fermentation medium consisted of: 20 g glucose, 2 g yeast extract, 1.5 g ferrous sulfate, 0.3 g 1 / 2 MS medium, and 1 L water. The fermentation liquid was inoculated into the substrate. After inoculation, the mycelium in the fruiting bottles / bags was cultured at 28–30℃ for 30 days until the mycelium had fully colonized the bottles / bags. The cultivation substrate formula was: 40% sawdust, 20% wheat grains, 10% wheat bran, 20% cottonseed hulls, 10% corn flour, 0.1% gypsum powder, and 0.1% potassium dihydrogen phosphate. Equal volumes of red soil and peat moss were mixed, and water was added to achieve a moisture content of 55%. The casing layer was set to a thickness of 1.5 cm. After casing, a 0.2% potassium dihydrogen phosphate solution was sprayed for mushroom cultivation management, and the results were recorded. (See attached table for details.) Figure 8 .

[0042] Figure 8 The left-hand diagram shows the primordium induction and fruiting body growth in this embodiment, while the right-hand diagram shows the fruiting body at harvest time.

[0043] Process records show that it takes 2–3 days for the mycelium of *Boletus sinensis* to fully colonize the casing layer, and 3–4 days for primordia formation. A relatively large number of primordia (7–10) are observed, with a size of 2–4 cm. The casing layer surface is free of contamination by other microorganisms. Harvesting can begin 13–15 days after casing. At harvest, the cap diameter is 6–8 cm, the stipe length is 8–12 cm, and the stipe diameter is 2–4 cm. The yield per bottle / bag is 100–120 g, with a biological conversion efficiency of 31.75–38.10%. This method features a short primordia formation time, a large number of primordia, and rapid development of effective primordia into fruiting bodies, resulting in high fruiting body yield and high biological conversion efficiency, making it suitable for industrial-scale production. Example 3

[0044] The *Boletus chinensis* strain JSJ-Bx1 was activated and inoculated into B1 medium for 5 days at 30–35℃. The B1 medium composition was: 20 g glucose, 2 g yeast extract, 1.5 g ferrous sulfate, 0.3 g 1 / 2 MS medium, 20 g agar, and 1 L water. Half a dish of activated mycelium that had fully colonized the plates was then chopped and subjected to liquid fermentation at 28–30℃ in the dark for 7 days. The liquid fermentation medium consisted of: 20 g glucose, 2 g yeast extract, 1.5 g ferrous sulfate, 0.3 g 1 / 2 MS medium, and 1 L water. The fermentation liquid was inoculated into the substrate. After inoculation, the mycelium in the fruiting bottles / bags was cultured at 28–30℃ for 30 days until the mycelium had fully colonized the bottles / bags. The cultivation substrate formula was: 40% sawdust, 20% wheat grains, 10% wheat bran, 20% cottonseed hulls, 10% corn flour, 0.1% gypsum powder, and 0.1% potassium dihydrogen phosphate. Equal volumes of red soil and peat moss were mixed, and water was added to achieve a moisture content of 60%. The casing layer was set to a thickness of 1.5 cm. After casing, a 0.1% potassium dihydrogen phosphate solution was sprayed for mushroom cultivation management, and the results were recorded. (See attached table for details.) Figure 9 .

[0045] Figure 9 The left-hand diagram shows the primordium induction and fruiting body growth in this embodiment, while the right-hand diagram shows the fruiting body at harvest time.

[0046] Process records showed that it took 3–4 days for the mycelium of *Boletus sinensis* to fully colonize the casing layer, and 3–5 days for primordia formation. The number of primordia was relatively small, at 3–5, and their size was 1–3 cm. A small amount of other microorganisms contaminated the surface of the casing layer. At harvest, the cap diameter was 5–7 cm, the stipe length was 8–10 cm, and the stipe diameter was 2–3 cm. The yield per bottle / bag was 90–110 g, and the biological conversion efficiency was 28.57–34.92%. This method has a slow primordia formation time and a relatively small number of primordia formed; it could be optimized for industrialized cultivation.

Claims

1. A species of *Boletus sinensis* that induces saprophytic growth (… Buchwaldoboletus xylophilus A method for rapid primordium formation, characterized in that, Includes the following steps: The first step involved activating the *Boletus saprophyticus* strain JSJ-Bx1 and inoculating it into B1 medium for dark incubation for 5 days at 30–35°C. The *Boletus saprophyticus* strain JSJ-Bx1 has the accession number CGMCC No. 42007 and is deposited at the China General Microbiological Culture Collection Center. The B1 medium consisted of the following components: 10–30 g glucose, 2–6 g yeast extract, 1–3 g ferrous sulfate, 0.1–0.3 g 1 / 2 MS medium, 18–20 g agar, and 1 L water. The second step is to carry out liquid fermentation culture on the activated mycelium that is about to cover the plate. The culture temperature is 28–30℃ and the culture is carried out in the dark for 7 days to obtain the fermentation liquid. The third step is to inoculate the fermentation liquid into the cultivation medium and incubate it in the dark for 30 days until the mycelium fills the bottle / bag. The incubation temperature is 28–30℃. The fourth step is to cover the bottles / bags filled with mycelium with soil, and then cover the surface with non-woven fabric as a protective layer for mycelium cultivation in the soil layer. Cultivate for 3-5 days to obtain the mushroom-growing bottles / bags with soil covering. The fifth step is to place the mushroom bottles / bags covered with soil into a mushroom house at 26–28℃ for light cultivation. During cultivation, spray with potassium dihydrogen phosphate solution to promote mycelial growth and induce primordia formation.

2. The method for inducing *Boletus sinensis* (a type of fungus) according to claim 1. Buchwaldoboletus xylophilus A method for rapid primordium formation, characterized in that, In the fourth step above, the nonwoven fabric has a density of 20 g / m². 2 The non-woven fabric is covered with soil with a moisture content of 50–60% and a thickness of 1–3 cm.

3. A method for inducing *Boletus sinensis* (a type of fungus) according to claim 1 or 2. Buchwaldoboletus xylophilus A method for rapid primordium formation, characterized in that, The mass concentration of the potassium dihydrogen phosphate solution used in step 5 above is 0.05–0.3%.

4. The method for inducing *Boletus sinensis* (a type of fungus) according to claim 3. Buchwaldoboletus xylophilus A method for rapid primordium formation, characterized in that, The covering material has a moisture content of 55%, a covering thickness of 1.5 cm, and a potassium dihydrogen phosphate solution concentration of 0.2%.

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