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OfflineCliftonGK1
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Does this agar plate growth look odd?
    #19347820 - 12/30/13 06:33 PM (10 years, 1 month ago)

I started a few plates a couple weeks ago from an old spore syringe and got a nice clean single start on one plate. After some days of letting it grow, I sectioned it out to new plates.  They took off relatively quickly (this is 4 days post transfer to new plates) but the odd, almost hyphal knotting look to the mycelium is bugging me. 
Is it normal to see that sort of knotting on an agar plate?

Growth conditions are pretty warm (75 - 76F) and this plate has been kept in a dark closet with the exception of my examinations, which total less than 5 minutes over it's entire growth.  It's growing on MEAY, if that makes any difference (although the PDYA plates look similar, with that ring of consolidated knotting near the original wedge.)

The transferred wedge was from a plate which didn't exhibit that knotting feature.  Transfers were done with a new sterile scalpel blade in an SAB.  Plates were wiped with 70% isopropyl prior to introduction to the SAB, as were all other materials.
The wedge didn't exhibit the knotting feature until after 2 days growth past the circumference where it now shows; so the mycelium grows as you see at the outer perimeter, in a lightly rhizomorphic appearance, and then seems to consolidate/knot after 36 - 48 hours afterwards.

Being an old-school microbiologist, I did the first thing any old-schooler would do; I sac'd one transfer plate to open it up and give it a sniff test.  (Us old-schoolers can ID lots of contaminants by appearance and smell.)  But this doesn't smell like anything out of the ordinary for a mushroom mycelium... Just the light yeasty smell of the agar plate, and the fresh earthy smell of the mycelium.  No sweetness or sourness to it at all.



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OfflineGod Stamet
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Re: Does this agar plate growth look odd? [Re: CliftonGK1]
    #19347863 - 12/30/13 06:43 PM (10 years, 1 month ago)

I would do a transfer from the outer edge where it looks healthy, the inside doesn't look good. the outside looks alright though so you should be able to clean it out. good luck


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OfflineCliftonGK1
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Re: Does this agar plate growth look odd? [Re: God Stamet]
    #19353961 - 01/01/14 08:26 AM (10 years, 1 month ago)

Quote:

God Stamet said:
I would do a transfer from the outer edge where it looks healthy, the inside doesn't look good. the outside looks alright though so you should be able to clean it out. good luck




So I did do a transfer from the outer edge of a couple plates onto new ones to see if the same thing happens.  I have a feeling that in a couple days I'll be seeing this again.
I left a plate with those odd knotting features sitting at daylight conditions, and it started to fruit. 
Now, I don't wrap my dishes with parafilm, I just keep them in a storage tote so there's no airflow around them.  This might be a product of not keeping a high CO2 concentration during the mycelial growth on the plate?



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OfflineGod Stamet
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Re: Does this agar plate growth look odd? [Re: CliftonGK1]
    #19354459 - 01/01/14 11:57 AM (10 years, 1 month ago)

if I didn't wrap my dishes with parafilm I would wrap them with something else for sure. saran wrap, scotch tape, something. even in a tote there's a small amount of air exchange which puts your dishes at risk. especially if you're like me and own pets. just keep cutting healthy growth away from bad growth and hopefully you get it cleaned out. you'll know for sure in a couple more transfers. otherwise start from scratch and lesson learned. good luck


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I have caught the Mycology bug! Not just here for the ride, i'm here for life.

How I do my bulk substrate

just another mushroom capsule TEK


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InvisiblePastywhyteMDiscord
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Re: Does this agar plate growth look odd? [Re: God Stamet]
    #19354594 - 01/01/14 12:45 PM (10 years, 1 month ago)

:whathesaid: If I'm feeling lazy an unused ziplock bag works good.


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OfflineCliftonGK1
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Re: Does this agar plate growth look odd? [Re: Pastywhyte]
    #19377590 - 01/06/14 02:55 PM (10 years, 1 month ago)

The transferred plate I left in similar conditions (no parafilm, no baggie, no nothin') is again doing the same knotting thing after a few days of strong mycelial growth.
I am not currently concerned, because from another plate of similar appearance, I let it go for another 3 days and all those knots started to pin.
So here's what I'm doing with a couple of those guys:  I've transferred out 4 pins taken from 3 plates which I let go to the point of pinning up.  Moved those cut pins to individual MEAY plates and this time I DID seal them up.  Didn't have parafilm, ended up using electrical tape.  Kinda chump looking and I hope that it doesn't mess anything up, but it's an experiment, so live and learn, right?
Anyhow, here's one of the transferred pins 24 hours post re-plating; looking happy and there's already some mycelial fuzz reaching out down to the new agar surface.


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OfflinecronicrFacebook
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Re: Does this agar plate growth look odd? [Re: CliftonGK1]
    #19377734 - 01/06/14 03:23 PM (10 years, 1 month ago)

:cool:


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OfflineCliftonGK1
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Re: Does this agar plate growth look odd? [Re: cronicr]
    #19390013 - 01/08/14 06:56 PM (10 years, 1 month ago)

72 hours post transfer and here's how things look:
All 4 transferred pins.  You can see that from origin plate #1 (left) the growth is fine and wispy and barely covers half the diameter of the growth seen in the #2a/b plates.  But both the #2a/b plates are showing that ring of knotting/spotting I've been trying to eliminate (although those spots *do* later turn out to be pins.)  The #3 plate (right) is sort of the midground on growth; better than #1, worse than #2a/b, and still has those spots, which the #1 plate is currently lacking.


Any ideas? 
Experienced cultivators:  I've left the origin plates for well past their prime.  They've grown scraggly ugly fruits which have since turned to gloop, yet those spots have either pinned or just stayed as spots.  There is no other evidence of any contamination.  Do you think the spots appear to be a contam, or just knotting?
Is knotting common on agar, or might it be something indicative of a specific characteristic I'm now isolating?

Here's a closer pic of the nearly concentric growth on plate #2b, and you can see that there's a pretty distinct ring at which those knotted/spotty bits start.  I can't seem to correlate them to any particular condition; I sealed my plates this time and temperature in my apartment is consistent.  Aside from the photo op and a brief check once a day, they're kept in a closet which doesn't get opened.


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InvisiblebodhisattaMDiscordReddit
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Re: Does this agar plate growth look odd? [Re: CliftonGK1]
    #19390024 - 01/08/14 07:02 PM (10 years, 1 month ago)


Knotting happens a lot on agar if you wait long enough, I would clone these but these dishes are already from a clone, If you do MS to a dish and let it grow some shrooms you could clone those rather than finding an isolate. I use the shrooms that grow on the agar as source for spores since they're going to be cleaner and easier to keep clean for printing than ones grown in a fruiting chamber.


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OfflinePaulsage
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Re: Does this agar plate growth look odd? [Re: bodhisatta]
    #19390504 - 01/08/14 08:37 PM (10 years, 1 month ago)

*awesome* thread - especially given that i'm about to drop some MS onto an agar plate in about 45 mins...

thx for all this...  :kaneclap:


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OfflineGod Stamet
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Re: Does this agar plate growth look odd? [Re: Paulsage]
    #19393149 - 01/09/14 11:05 AM (10 years, 1 month ago)

fruit it out. it'll answer all your questions. good luck


--------------------
I have caught the Mycology bug! Not just here for the ride, i'm here for life.

How I do my bulk substrate

just another mushroom capsule TEK


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OfflineCliftonGK1
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Re: Does this agar plate growth look odd? [Re: God Stamet]
    #19393271 - 01/09/14 11:28 AM (10 years, 1 month ago)

Quote:

God Stamet said:
fruit it out. it'll answer all your questions. good luck




That's the plan, man!  Working these guys up to one more round of plates, and then into grain jars.:thumbup:


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