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OfflinePBJ710
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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27056610 - 11/25/20 07:56 AM (2 months, 30 days ago)

A few plates look like they're having some issues, but 1 and 6 are looking the best so far (maybe 4 as well, but can't tell from pic).  I would give it another day or two so the culture can better show what you're working with.


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Re: Can you help me evaluating these Agar dishes? [Re: PBJ710]
    #27057018 - 11/25/20 01:45 PM (2 months, 29 days ago)

IMO give it a 3-4 days and take a transfer a cm or so on the inside edge.  I think what you are seeing are metabolites usually from the myc fighting a bacteria.


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Re: Can you help me evaluating these Agar dishes? [Re: IdiotCircusBoy]
    #27057933 - 11/26/20 01:00 AM (2 months, 29 days ago)

@PBJ710 @IdiotCircusBoy Thank´s a lot for the advice. I will let it go for a few days and take a transfer.
Right now, it looks like it is #6 that is the only one without mold coming out from the transferred lump of Agar.


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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27063982 - 11/30/20 07:00 AM (2 months, 25 days ago)

Hi again,

The dishes have evolved, and it is clear that I am transferring contamination from the gen. 1 dishes (from spores) to gen. 2 dishes.
If you remember, the Agar on the gen. 1 dishes was PC incorrect.

I post pics of all the dishes again, just in the interest of science :-)

Do you think #6 will be useful for transfer?

What do you think is the source of contamination? Gen. 1 Agar, Spores, or airborne in SAB?









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OfflinePBJ710
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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27064176 - 11/30/20 10:42 AM (2 months, 24 days ago)

Looks like 6 is the only candidate at this point.  I would take a couple tiny transfers from the leading edge and hopefully they will be clean(er).  It's entirely possible that the mold is still riding along with the mycelium at this point.  Personally I would transfer any good looking transfers 1 more time (assuming growth is orderly and not just a bunch of random fuzz) before expanding them, but it could probably be done directly from those T2 plates if they look really nice.


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Re: Can you help me evaluating these Agar dishes? [Re: PBJ710]
    #27064241 - 11/30/20 11:36 AM (2 months, 24 days ago)

Quote:

PBJ710 said:
Looks like 6 is the only candidate at this point.  I would take a couple tiny transfers from the leading edge and hopefully they will be clean(er).  It's entirely possible that the mold is still riding along with the mycelium at this point.  Personally I would transfer any good looking transfers 1 more time (assuming growth is orderly and not just a bunch of random fuzz) before expanding them, but it could probably be done directly from those T2 plates if they look really nice.




Thank´s a lot:-)

I will do so as soon as I get some spare time.  I´ll be back with the results when they have colonized a bit.

Best regards

N


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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27064411 - 11/30/20 01:29 PM (2 months, 24 days ago)

I'm def no contamination expert but I think green is typically Trichoderma and, as I understand it, you can't transfer your way out of Trich.  If 6 isn't green then you may have a contender.


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Offlineneonero
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Re: Can you help me evaluating these Agar dishes? [Re: IdiotCircusBoy]
    #27064536 - 11/30/20 02:33 PM (2 months, 24 days ago)

Quote:

IdiotCircusBoy said:
I'm def no contamination expert but I think green is typically Trichoderma and, as I understand it, you can't transfer your way out of Trich.  If 6 isn't green then you may have a contender.




Hmm. #6 is also a little green I think. I can try one transfer and see.


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OfflinePBJ710
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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27064964 - 11/30/20 06:14 PM (2 months, 24 days ago)

I was trying to be optimistic and hoping that it is just bruising.  We should be able to tell from the T2 plates how it's doing in regard to contams.  If they show bad stuff, I would start some fresh plates instead of fighting this out trying to clean it up.


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Re: Can you help me evaluating these Agar dishes? [Re: PBJ710]
    #27065615 - 12/01/20 01:12 AM (2 months, 24 days ago)

Quote:

PBJ710 said:
I was trying to be optimistic and hoping that it is just bruising.  We should be able to tell from the T2 plates how it's doing in regard to contams.  If they show bad stuff, I would start some fresh plates instead of fighting this out trying to clean it up.




:thumbup: :smile:


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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27071517 - 12/04/20 12:03 PM (2 months, 20 days ago)

Hi,

It turned out that #6 also developed green mold before I could transfer, so I have made up new dishes and inoculated with spores.

I´ll post pics when they start to grow.

Best

Neo


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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27071714 - 12/04/20 02:01 PM (2 months, 20 days ago)

Because I'm learning agar right now, could you please tell me how you think on your 1st attempt you got so many mycelium spots everywhere?

When you used the loop did you drag it all over your entire dish?  I ask because the amount of isolated(?) mycelium clumps from edge to edge of your dish confuse me.  I'm wondering if this is what others were reacting to at first.  I thought you only put a transfer in the middle, or drag your loop a few times.  Does this make sense?  thanks


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Re: Can you help me evaluating these Agar dishes? [Re: rashomon]
    #27073089 - 12/05/20 10:09 AM (2 months, 19 days ago)

Quote:

rashomon said:
Because I'm learning agar right now, could you please tell me how you think on your 1st attempt you got so many mycelium spots everywhere?

When you used the loop did you drag it all over your entire dish?  I ask because the amount of isolated(?) mycelium clumps from edge to edge of your dish confuse me.  I'm wondering if this is what others were reacting to at first.  I thought you only put a transfer in the middle, or drag your loop a few times.  Does this make sense?  thanks




Hi,

I used this streaking technique https://www.shroomery.org/forums/showflat.php/Number/24336286#24336286


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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27073544 - 12/05/20 03:08 PM (2 months, 19 days ago)

Quote:

neonero said:


Hi,

I used this streaking technique https://www.shroomery.org/forums/showflat.php/Number/24336286#24336286




I had no idea this was a thing!!  Thanks for explaining!


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Re: Can you help me evaluating these Agar dishes? [Re: rashomon]
    #27082061 - 12/10/20 02:07 PM (2 months, 14 days ago)

Hi again,

OK, so I started over with spores. This time, the making of Agar and the pour was fine. I took the alu foil with the print inside the SAB after washing it with alcohol. Scraped a little bit loose, and holding the foil over the dish, shaking a little bit on the agar plate.

The pics were taken after 2-3 days. It really does not look promising? What do you think?





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OfflinePBJ710
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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27082237 - 12/10/20 03:46 PM (2 months, 14 days ago)

Looks like a little tuft of mycelium on #3 that you should be able to use if that's not bacteria behind it - it may take a few days for them to show more colonies.


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Re: Can you help me evaluating these Agar dishes? [Re: PBJ710]
    #27082409 - 12/10/20 05:06 PM (2 months, 14 days ago)

#3 Maybe but its surrounded by something not kosher.  Did alcohol get on the spores or the plate?  I've not heard of the sprinkle on technique so am a little unsure about it.  The trick with a SAB is to not let stuff get above what you are working on.  So using a sterilized inoculation loop to swab the spores and swipe the plate accomplishes this because you're coming in from the side.  Your also using a streak technique so that spores are spread out on the plate.

Check out Bod's video on Spore Print to Agar.
https://www.shroomery.org/forums/showflat.php/Number/24337011#24337011


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Edited by IdiotCircusBoy (12/10/20 05:08 PM)


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Re: Can you help me evaluating these Agar dishes? [Re: IdiotCircusBoy]
    #27083214 - 12/11/20 01:10 AM (2 months, 14 days ago)

Quote:

IdiotCircusBoy said:
#3 Maybe but its surrounded by something not kosher.  Did alcohol get on the spores or the plate?  I've not heard of the sprinkle on technique so am a little unsure about it.  The trick with a SAB is to not let stuff get above what you are working on.  So using a sterilized inoculation loop to swab the spores and swipe the plate accomplishes this because you're coming in from the side.  Your also using a streak technique so that spores are spread out on the plate.

Check out Bod's video on Spore Print to Agar.
https://www.shroomery.org/forums/showflat.php/Number/24337011#24337011



Thank´s for the advice. I have used Bod´s tek in the past, but this time I did the sprinkle. I found it on youtube, but what I did not think of, was that it was used with a flow hood. I will make a couple of new ones with streaking this weekend.


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OfflineGrinchGrower
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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27083310 - 12/11/20 02:39 AM (2 months, 14 days ago)

All the growth underneath the black specks looks like bacteria.

The print may just be really dirty...

Someone suggested hot pouring a plate to get the myc racing away for a cleaner transfer. Might be worth a shot on #3.


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Re: Can you help me evaluating these Agar dishes? [Re: neonero]
    #27084101 - 12/11/20 04:49 PM (2 months, 13 days ago)

Did you use alcohol on the spore print?

Quote:


I took the alu foil with the print inside the SAB after washing it with alcohol.





No alcohol on the spores or plate.


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