Shiitake spawn preparation: Shiitake, Lentinula edodes, is a saprophyte and white rot fungus which feeds on deal oak tree species in nature. In its life cycle, each basidium under the gill of a fruit-body produces 4 basidio-spores which sporulate under a certain condition to become a primary mycelium and then a secondary mycelium by fusion between compatible mycelia.
Shiitake spawn is a medium, colonized by the secondary mycelia. The second mycelia will eventually form fruit-bodies.
Factor regarding mycelial growth of shiitake: Temperature: 21-27oC, RH%: 95-100; Duration (days): 35-70; CO2 (ppm): >10.000; Fresh air exchange: 0-1/hour; Light (lux): 50-100.
Shiitake spawn can be made using highly quality of fresh shiitake as the inoculum. A fruit-body is cut into 2 pieces vertically, and then a small piece is taken from a cut surface of the fruit-body and transferred to PDA media. Then this is incubated. The strain is transferred to PDA media in several times to get pure culture. Then the strain is incubated in a spawn substrate such as sawdust, wood plugs or grains. When the acquire sufficient quantities, the spawn is ready for distribution.
1. Mycelial transfer from PDA in tube (strain) to PDA in petri dish
Material: strain (mother culture in tube); PDA media; clean bench equipped with air filter, burner, alcohol 70%; transferring tools; UV lamp; autoclave, flask, petri dishes, scale, tin foil, incubator…
Prepare PDA media in : http://sietalab.com/study-the-growth-of-mushroom-mycelium-on-various-agar-media/
Transferring: Sterilize the tool until it become red-hot in the flame (burner), one hand grabs tube (plate) and other hand grabs tool, cool the tool by touching it on the media, hang the tube mouth on the flame for while; open the cap with a little finger; cut a sample form mother culture. Tube mouths and tools are flamed on the alcohol lamp before each opening and action; slightly open the petri plate and place the sample to the center of the PDA media, close the cap; wrap up the complete dishes with plastic; Record the required information such as date and strain name.
Problems can occur during this stage and the solutions of those problems are mentioned below:
2. Mycelial transfer from PDA petri dish to spawn substrate (sawdust)
Material: mother culture in petri dish; substrate materials; container, sterilizer, mixer and spawning room; incubation room.
Fill the heat resisting container (bottles, bags) with a substrate mixture, 850cc bottle is usually filled with 550cc substrate. Compact the surface of the substrate and make the inoculation hole (1.2-2cm). Close the cap and remove substrate particles around the bottle mouth; sterilize the bag just after filling at 121 oC for 60 minutes. After cooling, transfer the mycelia from incubated mother culture in petri dish into inoculation hole of substrate in the bottles/bags. Record needed information. These inoculated substrates are kept in room temperature.
Problems can occur during this stage and the solutions of those problems are mentioned below