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Post Number: 9569
|Posted on Saturday, December 20, 2003 - 05:57 pm:||
Rye Grass Seed in Autoclavable Bag with Injector Port
Below you will find instructions for the preparation of rye grass seed in Autoclavable Spawn Bags. The bags we are using for this demonstration contain filter patches and our new self-healing injector site. For these instructions we will be filling 3 bags and sterilizing them in a #915 All-American Pressure Cooker.
1. Fill 3 Autoclavable Spawn Bags with 500 gm of Rye Grass Seed.
2. Add 500 ml of water to each bag.
3. Mix well.
4. Fold over top of bags and hold closed with a clothespin.
Flip bags over but keep opening at top like in photo below.
Let sit like this for 8-10 hours or overnight. Rotate bags back to the normal upright position and fold the top down.
5. Now load 3 bags into a #915 All-American Pressure Cooker.
Add 3000 ml water to cooker.
6. Place extra bottom rack on top of the bags. This prevents the bags from blocking the vents of the cooker.
7. Put the lid on your cooker and secure. Turn on the heat to the range and set on high.
8. Let the steam vent until the pressure gauge is near 5 pounds pressure. This will ensure you that all air has been forced out of the unit.
Place the rocker on the vent pipe and turn down the heat to medium-high. This will vary with your particular range.
9. Pressure will slowly build until it reaches 15 pounds and then it will escape from the rocker. At this time turn the heat down to medium or so, just enough to maintain the pressure without making too much noise.
Once it reaches 15 pounds hold it there for 2 hours then turn off heat. Do not move the cooker. To many, 2 hours is overkill but I prefer double sterilization times for grain products
10. Now is a good time to clean the work area where the bags are unloaded from the cooker.
When the pressure has dropped to less than 5 pounds, the cooker can be moved, with care, to rest until opened.
11. This demonstration will be done in front of a HEPA (High Efficiency Particulate Air) filter system. A good still-air chamber or glove box will also work but contamination is more likely. It can be done.
Make sure you have washed your hands before proceeding.
Remove the rocker weight and open the pressure cooker.
12. Remove the 3 bags and let cool overnight or until less than 80 degrees (F).
13. Once cool the bags can be sealed with an impulse sealer.
14. The bags are now ready for inoculation.
Using your culture syringe, gently insert the needle through the small injector disk. Squirt approximately 2-4 ccs of spore solution into the rye grass seed. Slowly remove needle and the injection site will seal itself (If you are using bags without injector sites, insert needle through the side of the bag and tape over the hole once needle is removed).
Care for bags and store at proper incubation temps as species requires.
Post Number: 60
|Posted on Monday, December 22, 2003 - 02:22 am:||
How quickly do these colonize? Also, where can I get bags like that really cheap? (under $1/each?)
Post Number: 9696
|Posted on Monday, December 22, 2003 - 02:47 am:||
i suppose i should have mentioned that rye grass seed is used for sclerotia-growing species like mexicana.
they colonize pretty fast, you can't 'shake'
but you can 'knead' the bags to
spred mycellia around to help.
check our sponsors list for bags,
several carry them.
(Message edited by admin on December 22, 2003)