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OfflinePTreeDish
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Help Needed IDing Growths on Agar
    #26493761 - 02/19/20 05:22 PM (3 years, 11 months ago)

Growing 2 diff strains of cubensis from spore prints. It's been about a week at ~45% humidity 60-75 F incubation. Growth seems slower than what I've read I should expect. Starting to wonder if there are any viable prospects here...

Can I get some help on these? I left more specifics observations and questions next to each photo (click for larger size).

Would be MUCH appreciated.  :whiteflag:

1. This ones easy - I see some black web contam in top right. No other growth.



2. There's mold in the center, but what about the colony in lower left? Worth transferring?



3. Looks like the round growth has sectors, but it's kind of shiny in appearance. Also note growth on rim. Not sure if anything is good here?



4. Might be the best plate, but the growth is not symmetrical and I see little ridges or folds on the colony. What do you think?



5. Looks promising?


6. Look good? Why so slow after 1 week?


7. A fetus growth, very symmetrical growth and a few spore colonies. Any concerns?


8. Colonizing along the edge, next to a lighter whispy growth.


Edited by PTreeDish (02/19/20 05:24 PM)


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OfflineMr.Wizard
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Re: Help Needed IDing Growths on Agar [Re: PTreeDish]
    #26493830 - 02/19/20 06:00 PM (3 years, 11 months ago)

Spore germination can take up to 30 days, just have to hurry up and wait.

5 is the only one that looks like mine did. I have different varieties though. I for sure see contams, but on some of those I can't tell you if it's myc or not.


--------------------
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I am so happy and grateful that we get to live in joyful abundance, while things get better, and better.


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InvisibleInfiniteDreams
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Registered: 10/25/19
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Re: Help Needed IDing Growths on Agar [Re: PTreeDish]
    #26493841 - 02/19/20 06:05 PM (3 years, 11 months ago)

No, you aren't growing two different strains, you are likely growing thousands if not millions of different strains.  You chose two different varieties to start with.  Now the bad news...

:eww: #1 - #4 - chuck those yesterday :eww:

#5 - there you go, looks like MS germination

The rest I'm not so sure of given your front runners...  so guesses here but 6 and 8 are likely bad news.  For #7 there are 3 good colonies to the left but aggressive contamination in two spots already started so not good hopes for that one

So, getting focused, you have huge contamination issues, what is your basic approach?

Clean yourself, your environment, your materials.
Proceed with a focused an clean methodology.  If any of this is confusing or vague then take advantage of the resources here to learn.


Edited by InfiniteDreams (02/19/20 06:24 PM)


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OfflinePTreeDish
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Re: Help Needed IDing Growths on Agar [Re: InfiniteDreams]
    #26494227 - 02/19/20 11:21 PM (3 years, 11 months ago)

Quote:

InfiniteDreams said:
No, you aren't growing two different strains, you are likely growing thousands if not millions of different strains.  You chose two different varieties to start with.  Now the bad news...

:eww: #1 - #4 - chuck those yesterday :eww:

#5 - there you go, looks like MS germination

The rest I'm not so sure of given your front runners...  so guesses here but 6 and 8 are likely bad news.  For #7 there are 3 good colonies to the left but aggressive contamination in two spots already started so not good hopes for that one

So, getting focused, you have huge contamination issues, what is your basic approach?

Clean yourself, your environment, your materials.
Proceed with a focused an clean methodology.  If any of this is confusing or vague then take advantage of the resources here to learn.




Appreciate the correction on strain vs. varieties. I'm still learning the right terminology and for some reason, those two I mix-up all the time.

Re: 1-4. Toss 'em OK. You don't see anything that resembles MS? I could use a good resource that shows various examples of healthy MS growth in various stages and from various sources (spores vs transfers, etc). Would be really helpful in learning how to separate the wheat from the chaff.


> So, getting focused, you have huge contamination issues, what is your basic approach?
Well, I poured 50 plates in this batch and the only ones that show contam are the ones I cultured, ostensibly ruling out anything related to the process of preparing and pouring the agar.

I also used two spore prints from a vendor and found the spores very difficult to scrape off. One was on a small piece of white card stock and I found it much harder to get good scrapings compared to the foil.

I also (don't throw anything at me please), cooled off my scalpel by wiping it in between a soaked 70% IPA paper towel. At the time, this seemed like a good idea right as I've been having some issues with carbon flaking off the blade into my first few plates. Of course, after I did this, I read that I was probably just smearing contams all over my blade after flaming it and then scraping the contams into the print and onto the dish.

Quote:

Clean yourself, your environment, your materials.
Proceed with a focused an clean methodology.  If any of this is confusing or vague then take advantage of the resources here to learn.




A little presumptive and preachy, but I don't disagree with the sentiment. I'm learning and my mistakes I've accepted are part of the process. I humble myself before internet strangers in judgement to teach and guide me towards incremental improvement.

So do you think I should try and transfer those 3 colonies in #7 to new plates before its too late? I'll also be sure to post an update in a week or two with new pics.


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OfflinePTreeDish
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Re: Help Needed IDing Growths on Agar [Re: Mr.Wizard]
    #26494236 - 02/19/20 11:32 PM (3 years, 11 months ago)

30 days OK. I had 7 days stuck in my head which I swore I read in another post. Good to know though. :thumbup:


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OfflineMr.Wizard
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Re: Help Needed IDing Growths on Agar [Re: PTreeDish]
    #26494266 - 02/20/20 12:19 AM (3 years, 11 months ago)

Use your #5 as a reference from now on. There are literally thousands of pictures here of spores on agar if you want to find what others look like.

Infinite Dreams wasn't implying you're a dirty person, just that the cleaner you are the better. Like making sure someone has turned their whatever electronic off and on again to try and fix it first. You obviously have contams, so now you need to find the vector.

Here's some RustyWhyte spores from a syringe on agar:



Here's some first transfer plates:



--------------------
Tricks to the search bar: https://www.shroomery.org/forums/showflat.php/Number/24270830

Where to Start: https://www.shroomery.org/forums/showflat.php/Number/24420178/fpart/52#27623666

My easy to see modified no-pour: https://www.shroomery.org/forums/showflat.php/Number/26467702

I am so happy and grateful that we get to live in joyful abundance, while things get better, and better.


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OfflineMLPismyOPSEC
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Re: Help Needed IDing Growths on Agar [Re: Mr.Wizard]
    #26494594 - 02/20/20 07:52 AM (3 years, 11 months ago)

5 is the only one that looks good to me, everything else is contam'd. Don't throw out those plates just yet though, the contam may not spread and you could still potentially get good myc from it. But focus on 5, she's lookin good. First germination plates always take a while, but in a week you should have some good spots to transfer from.


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InvisibleInfiniteDreams
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Re: Help Needed IDing Growths on Agar [Re: MLPismyOPSEC]
    #26494623 - 02/20/20 08:09 AM (3 years, 11 months ago)

Quote:

MLPismyOPSEC said:
Don't throw out those plates just yet though, the contam may not spread and you could still potentially get good myc from it.





I disagree.  Get rid of those plates.  Such an aggressive contamination should not be fostered in anyway.


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OfflineSvetaketu
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Re: Help Needed IDing Growths on Agar [Re: InfiniteDreams]
    #26494671 - 02/20/20 08:36 AM (3 years, 11 months ago)

I say throw out everything but #5 and #7.

#5 looks clean, so prioritize that one. #7 looks like 3 clean germination spots on the bottom left and 2 spots of bacteria on the right. Bacteria grows slow, so just transfer the clean parts away from it.

FWIW, you are looking for white, fuzzy, organized growth. Here are some clean germ plates examples;

Anything with that filmy/wet appearance is some kind of bacteria. Anything fuzzy with color means it's a mold, and it's already producing spores, so be cautious around those.

Also, if you really want a place to cool your scalpel, you can cool it in the clean agar dish your about to scrape spores onto.

Good luck! :super:


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LAGM2021


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OfflinePTreeDish
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Re: Help Needed IDing Growths on Agar [Re: Svetaketu]
    #26497419 - 02/21/20 05:57 PM (3 years, 11 months ago)

You folks are tremendous.

I went ahead and ditched all but #5 and #7. Did 1x T1 for #5 and 3x T1s for #7. Also went ahead and made a few new cultures from print. First time doing agar xfers. Hopefully I get something viable from these.

Keep trying to improve my sterile technique. The only thing I can't control (yet) is the cleanliness of the prints I'm using.


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OfflinePTreeDish
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Re: Help Needed IDing Growths on Agar [Re: PTreeDish] * 1
    #26500249 - 02/23/20 05:37 PM (3 years, 10 months ago)

Thought I'd post a little update here. Following your advice, I ditched all but #5 and #7.

Here's #5 now:



Growth continues and sectoring appears to becoming more visible. Growth is def. whispy / cottony in appearance. Note that there was another healthy colony on the edge that I transferred.

Here's #7 now:



3 good colonies next to 2 bacteria. Luckily, the bacteria hasn't really grown any more and I was able to take super small wedges from each colony and transfer them.

Now, I've got some nice fluffy healthy nuggets growing without contams. :smile: Yay.



I think not wiping the scalpel and cooling in the dish made a big difference in eliminating or cutting out contams on these transfers (my first ones btw).

I did a few more cultures from print and one already contam'd so I think there is indeed something funky with the prints.

:sun:


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