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1. Main Formulas for Master Spawn and Cultivation Spawn

(1) Sawdust Medium: Broad-leaved tree sawdust 78%, wheat bran 20%, sugar 1%, gypsum 1%; pH 6.5–7; moisture content 60%–65%.

(2) Corncob Medium: Corncob 78%, wheat bran 20%, sugar 1%, gypsum 1%; pH 6.5–7; moisture content 60%–65%.

2. Production Methods

(1) Selection of Spawn Bottles and Bags: Standard 500 mL spawn bottles are commonly used for master spawn; polypropylene bags (17 cm long, 33 cm wide, 0.004 cm thick) are generally used for cultivation spawn.

(2) Filling Bottles and Bags

① Filling Bottles: After washing and draining the spawn bottles, fill them with the culture medium up to the "shoulder" of the bottle; wipe the inner and outer walls clean and cap with a lid featuring an aeration hole.

② Filling Bags: Fill with culture medium to a height of 20 cm and attach the double-ring collar assembly.

(3) Sterilization Methods: There are two methods: high-pressure sterilization and atmospheric-pressure sterilization. For atmospheric-pressure sterilization, maintain 100°C for 10–15 hours; for high-pressure sterilization, maintain 1.5 MPa for 2 hours.

(4) Inoculation Methods

① Master Spawn Expansion Method (inoculation performed in a laminar flow hood; transferring from test tube to bottle or plastic bag): a. Disinfect hands and the exterior of the mother culture test tube; b. Light the alcohol lamp; c. Remove the cotton plug from the mother culture tube, flame-sterilize the tube opening and inoculation spoon over the alcohol lamp, and secure the mother culture; d. Remove the cotton plug from the spawn bottle, transfer two 1 cm² pieces of spawn into the bottle; e. Re-insert the cotton plug and affix a label. 

② Method for expanding cultivation spawn (inoculation inside a laminar flow hood; transferring from bottle to bottle or plastic bag): a. Sterilize the cotton plug of the master spawn bottle; b. Sterilize hands and the exterior of the master spawn bottle; c. Light the alcohol lamp; d. Remove the cotton plug, flame-sterilize the mouth of the master spawn bottle and the inoculation spoon, and secure the master spawn bottle in place; e. Remove the cotton plug from the spawn bottle, scrape away the old spawn clumps and mycelial mat from the surface, break up the spawn using the inoculation spoon, and transfer a full spoonful into the cultivation bag; f. Insert the cotton plug and affix a label.

(5) Mycelial incubation method: After inoculation, the spawn bags should ideally be incubated in a clean, dark room at a temperature of 20–25°C and an air humidity of 60%–70%, with ventilation performed 1–2 times daily. Additionally, the stacks should be turned and inspected every 7–10 days so that any issues can be addressed promptly. Under constant-temperature incubation at 22–25°C, a 500 mL secondary spawn bottle typically becomes fully colonized within 25–30 days.