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Offlinetarfoh
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First Time Agar Work - CONTAM CITY!!! help =(
    #14292121 - 04/14/11 07:35 PM (12 years, 10 months ago)

Hey everyone!  I have just gotten started with agar work and I have apparently taken on a difficult task.  What I am trying to do is clean up some wild cube prints that I received from europe.  I put some spores on four agar dishes, and all of them are showing signs of strange growth.  It is definitely contamination.  Two of the dishes have a disgusting liquid that oozes around when I move the dish, and some of the dishes have spots of cobweb and pre-sporulating trich.  Now in order to clean these up, I am going to attempt to transfer some healthy mycelium, if any at all, onto some new plates.


My question is how exactly do I do it without spreading more contamination?  Do I throw away the plates with the liquid that moves around when I lift the petri dish?  Is the goal to have my second transfer LESS contaminated or completely contam free?  I have researched and watched videos but doing all of this for the first time is rather nerve-wracking so I just want to know if anyone has any suggestions or experience with this.  Thanks ahead of time.


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OfflineDoDahDay
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Re: First Time Agar Work - CONTAM CITY!!! help =( [Re: tarfoh]
    #14292326 - 04/14/11 08:13 PM (12 years, 10 months ago)

Now, when I say this, please wait for a "trusted cultivator" to confirm before you take all of my advise as law.

First off you need to try to determine where the contam came from. How long did you ster. your dishes? How did you transfer the spores to the dishes. How were your spores stored? Were the dishes pre ster. in a plastic sleeve(plastic dishes) or autoclaved or ster glass? Look at the vectors of contam- cultivator, air, media, tools, inoculum, MDU(bugs or some sort)(from the GGMM).  Look, also, at the pattern from which you inoculated them and look at the contam that are present from the first to the last. Did it change, get worst or better? Could help you see where shit went wrong or got worst.

If you had some mycelium grow, and it has a healthy sector, in the dishes that are only showing bacteria contam(slimy liq or slimy spotting) you should be able to remove a piece of the myc and dip it in a weak solution of h2o2 and retrans. so long as you have corrected your vector of contam that has f'ed things in the first place. Still ify.

I believe ive heard of ,again dont take this as law til you here from the "trustees", using a weak salt water solution to make an inoculum to transfer spores to the media. If the spores have bact on them, the weak salt solution can help inhibit them if it is present on them.

If its trich, no sudden movements and retrans. Good luck.


--------------------
"I fart in your general direction!  Your mother was a hampster and your father smelt of elderberries!"


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OfflineJay Sav
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Registered: 03/14/11
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Re: First Time Agar Work - CONTAM CITY!!! help =( [Re: DoDahDay]
    #14297428 - 04/15/11 07:05 PM (12 years, 10 months ago)

Here is a great tek I read some time ago, if you havent read it, you definately should check it out.

The Agar Sandwich
http://www.shroomery.org/forums/showflat.php/Number/2823255#2823255

RR has a different method of doing this, involving hot agar being poured over the contaminated plate and then scraping off a minute fragment of mycelium when it grows through to the surface. He then transfers that tiny fragment to a new dish. I'll look it up and get back to you.


Edited by Jay Sav (04/15/11 07:12 PM)


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OfflineJay Sav
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Registered: 03/14/11
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Re: First Time Agar Work - CONTAM CITY!!! help =( [Re: tarfoh]
    #14297479 - 04/15/11 07:20 PM (12 years, 10 months ago)

Found it!

Quote:

RogerRabbit said:
One way I've found to beat those pesky contaminants is through agar pasteurization. Make up a fresh batch of agar. When it has cooled enough to pour into the dishes(+/- 120F-130F or 49C-55C), pour a layer right over the top of the dish with both healthy mycelium and the contaminants. Be sure to cover all the growth in the dish with a layer of hot agar. The mushroom mycelium, being a coherent network will usually survive the assault, but the contaminants will be neutralized for a few days. Watch the dish carefully. Usually within 48 to 60 hours, the mushroom mycelium will make its first appearance through the fresh layer of agar. Use an inoculating loop rather than your scalpel and immediately scrape a tiny bit of this mycelium off the surface of the top layer without taking any of the agar itself. This usually results in a very clean transfer of mushroom mycelium before the contaminant has had a chance to recover.




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