27 replies to this topic

[smegma]

Looking for opinions:

1. Cardboard glove box. (Clean, not "Sterile") Ousted + 70% ISO everywhere.
2. Chunk of "clean stem" ripped and dipped in 3% H2O2 fo 30 seconds. (Not clean stem), lots o fizzyness.
3. Into sterile blender jar w/ 100ml distilled. (Sterile)
3.4 "zap-zap" w/ blender 2-4sec. bursts. (Sterile)
4. Into sterile syringes. (Sterile)
5. noc 20 WBS jars. (Sterile)

What % do you think will NOT be contaminated?

Should I have shaken at innoculation or waited for recovery?

Water was dark ass blue and thik with myc.

[Hippie3]

are you taking the sample from inside the stem ?

[smegma]

are you taking the sample from inside the stem ?

Well, embarassingly I got a good chunk of the outside in there on accident. It did get the H2O2 treatment, but was not all core material.

You are thinking I am SOL?

I noc'd 20 qts. with it already... :(

If I could just get on qt. clean I could g2g or could their still be invisible contams and I should abort?

[I_am_me]

Its very hard to predict what will happen in such an uncontrolled environment.

You pretty much just have to prey they don't end up contammed.

[smegma]

Its very hard to predict what will happen in such an uncontrolled environment.

You pretty much just have to prey they don't end up contammed.

If I end up with a NICE arse White jar from this should I still be concerned with "invisible" bacteria? Or can I G2G it with relatively little concern? Could their be a latent contam that smacks me down way late in the game like during pinning?

Am I asking too much for 1 hour?

[smegma]

quote:
"3.4 "zap-zap" w/ blender 2-4sec. bursts. (Sterile)"

If you can afford a $500 stainless steel blender, then you can afford a flowhood. Simply use agar and forget all the hassle. If you have less than an eberbach blender, you can't claim 'sterile'. Nothing is sterile unless it has been through a PC cycle.

The jar, water, gasket, blades and metal disc cover were PC'd with a dome lid ring holding it in place tightly and wrapped in foil. PC'd for 30 min.

How is that not sterile?

[Lazlo]

Check this out! These were done in a H202 solution, and then on to a vinegar solution, drained well in open air, and into a PC'd 1 quart jar in gloove box. The bowl and jar cleaned as well as i could before loading into gloove box. Bacillus was the contam. (Wet Spot Bacteria) I left the jar in a pile of clothes about 2 months ago all noided out and found it putting the clothes away the other day. So I thought i'd give it a try to see what happened. There has been great feed back about the use of both for sanatary use. Do not fool around with bacterias or molds unless you use the right precautions and have experience.

Note: I'm not saying that you can be sloppy with a cloning tek using this method.

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[Lazlo]

Sorry! The point I left out is that if you use H202 for this purpose, follow it up with the vinegar solution to flush out the H202 some. H202 is horrible for mycelical growth. But followed up by the vinegar solution, it seems to help dramatically. There's been great feedback among communities on the use of both solutions for sanatization purposes. But H202 first. But like i said, don't try to get away with a sloppy cloning tek using this method.

[smegma]

Sorry! The point I let out is that if you use H202 for this purpose, follow it up with the vinegar solution to flush out the H202 some. H202 is horrible for mycelical growth. But followed up by the vinegar solution, it seems to help dramatically. There's been great feedback among communities on the use of both solutions for sanatization purposes. But H202 first.

I will try to get a clean core sample next time, BUT I will definitly try the Peroxide --> Vinegar for shits and giggles.

Thx for the info.

[Hippie3]

vinegar eh ?
you know,
back when i was first trying to develope my bleach dip,
i read a sanitizing formula that had bleach and vinegar
so i tried a mixture
and was bummed to see it made trich grow even faster
lol
but reading what you said just now
makes me wonder what part the vinegar
played, and what it could mean to healthy cakes.
where did you see more info on this ?

[Hippie3]

Vinegar is an acid. Trich loves acidic conditions.
Mycelium favors basic conditions

yes but
things aren't always as simple as first glance
i still wonder...

[Lazlo]

http://www.foreverhe...s/disenfect.asp

Check it out. I thought I might give it a try with seed to save my strain. The remainder of this wbs was from a 1qt totally eaten up with wet spot. I had it burried in some clothes and I guess it sort of choked it up. I wish i had taken pics of the 1qt. It was the absolute most horrible lookin shit you've ever seen. I didn't even think of documenting it with pics until this point. So far so good. The study showed that the use of H202 and vinegar were even more effective than bleach and some of the best cleaners in killing harmful bacterias AND molds.

[Lazlo]

Has anybody ever tried 35% food grade type of h202 that they talk about in the link above for myco work?

[sandman]

I as well have tried the blender/H202 clone method.

All I have to say is "Have fun cleaning those 20 Qt jars friend, cause my money is on contam if h202 and a blender was involved in any way shape or form"

Roger is correct when he says use a flow hood with agar instead of a blender. Its just the way it needs to be done. Or at least just cut the chunk up, and drop it in some karo water. Running a blender moves alot of air arround and inside the liquid being blended. Thats not a good thing for shizzle.

For wahtever reason I have found that actual mycelium is better for cloning than a peice of tissue. I have had zero success when it invlolves tissue from the actual shroom. I have had 99% success when I use mycelium. Thats just me, but its still a statistic.

Hopefully it works out for ya holmes, but I would be suprised.

[Lazlo]

Yeah. The blender tek isn't near as high % as just putting in Karo or petris. Remember, the more steps in a tek, the more chances of a contaminaton issue.

[Hippie3]

peroxide will not kill living molds, just the spores thereof.
bleach is much more effective
killing both spores and the mold colony itself.

[smegma]

I REALLY appreciate all of th input fellas. Due to my crampedquarters, I cannot use a flow hood. Agar in a glove box is a BIG time pain in tha ass. So I was hoping the blender TEK would give me a 10-20% success rate with the WBS jars so I could G2G from there.

Someday I will get a flow hood and be able to do agar, but till then I have to try this half assed shit considering how kick ass everyone here says cloning is.

THANK YOU ALL FOR YOUR INPUT.

Sorry if I come off as an ass sometimes. I don't mean it, the Xanax just wears off... :)

[smegma]

Vinegar is an acid. Trich loves acidic conditions.
Mycelium favors basic conditions.

If you'll cut a small piece of tissue from the very center of a stem, after tearing the stem lengthwise by hand(don't cut with a knife), you'll have a sterile tissue. Cut it with a scalpel that has been heated red hot. Cool the scalpel in the glovebox or in front of the flowhood and make the transfer. You don't need a chunk more than the size of a grain of rice. That makes it easy to cut a clean tissue from the center of the stem. This is my procedure and i don't ever use peroxide. It works every time.

I will assume that you then grow this out on agar?

Woult liquid culture be OK as a next step?

How abou right to grain?