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Ferather
Mycological



Registered: 03/19/15
Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather]
#25636531 - 11/24/18 02:52 PM (5 years, 3 months ago) |
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The peg has recovered, and now started growing out. Hopefully there is enough stored carbon to get a good speed. Otherwise it will take a while to start with, cellulose is relatively inert (non-reactive), slow to decay.
The additional phenols in the wood pellets should also help with speed, the pH is 7.5. Normally I would use starch based grain spawn with either WL-Tek recipe.
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Edited by Ferather (11/25/18 05:26 AM)
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25637557 - 11/25/18 05:39 AM (5 years, 3 months ago) |
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T-Gel, test recipe:
The Cubensis is continuing without the peg and nearly done, however there is now a spot of mold (probably from lid removal). I'm using a metal tea strainer so I have a good spread of leaf debris, I have taken high beam illuminated images.

Analysis:
Cubensis growth is surface based, although it's enzymes reach the bottom, in relation to position. Oxidation of the tea phenols is very apparent, phenol targeting enzymes are in use. The mold penetrates the agar, and appears to have germinated on debris.
No visible bacteria or yeast, nothing on the surface.
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Edit: Oops, sorry forgot to wait 24 hours, I'm tired. Sorry again.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25639666 - 11/26/18 07:58 AM (5 years, 3 months ago) |
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Peg to WL-Tek (lignicolous recipe):
The peg is working but slowly, I'm surprised it has any resources left.

This test sample will take a while.
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T-Gel (test recipe):
What I thought was mold is behaving strangely. The surface growth looks odd, and it's radial in the agar. The invader mycelium appears to be a white rot fungi, there is little to no browning present.
The conditions and media may not be ideal for the mold to sporulate.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25644260 - 11/28/18 06:05 AM (5 years, 3 months ago) |
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It doesn't look like Cubensis will conquer the invader mycelium, but perhaps it might cut it off. The orange previously seen is fading, possibly due to light or more O2 (oxygen).
As the mycelium pushes further, it gains more oxygen (container).
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Ferather
Mycological



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Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather] 1
#25646085 - 11/29/18 06:08 AM (5 years, 3 months ago) |
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WL-Tek test: 112g Water, 25g Paper Pellets, 12.5g Black Tea Leaves, 6g Ultra Fine CaCO3, 1g MG, 0.1g YN.
I started with 160g of water, it was then cooked down to 112g (~72% water), using a microwave. The water was added to the additives, mixed, then added to the pellets.
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T-Gel test: 100g Water, 2.4g Agar, 1.6g Black Tea, 0.04g MG, 0.02g YN | No sugar, or flour.
I transferred a wedge 12 hours ago, and have visible growth already. It was relocated it after the drop (hence the left patch).

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Now +6.5 hours from the previous image, and it's a personal record. Probably the nutrient combination (MG + YN) + phenols.
Edited by Ferather (11/29/18 04:02 PM)
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Ferather
Mycological



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Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather]
#25648927 - 11/30/18 03:50 PM (5 years, 2 months ago) |
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Growth is faster, thicker and very populated, carbon (phenol) utilization has increased with additional macro-micro nutrients. Current speed is faster than Tarragon oyster on T-Gel (plain, no flour) but enriched (MG @ 0.1g, no YN).
Even the mycelium left from the wedge relocation has regenerated and is growing out. The wedge now appears to be turning darker (phenols oxidizing).

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Edited by Ferather (11/30/18 04:26 PM)
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Ferather
Mycological



Registered: 03/19/15
Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather]
#25650986 - 12/01/18 04:05 PM (5 years, 2 months ago) |
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Working as expected, and it's fast, the added array of macro-micro nutrients combined with the carbon rich phenols makes a viable media. This suggests enzymes and-or reactions needed to decay and utilize the carbon source (phenols) have increased.
Note: There is almost no simple sugar (0.012%, 100g of T-Gel, from tea) and no starches.
I conclude Cubensis can and will target phenols for decay.

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02/12/2018 - 14:30 (+15 hours)
The WL-Tek test is working, the phenols are speeding up decay. However, more of the phenols are being spent on additional enzymes-other. Instead of just decaying phenols and growing out, more resources are being spent on decaying other materials (slower).
In addition, the soluble nutrients are more locked into the paper cellulose, and less free. The wedge is a little close to the lid, and producing condensate.

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Now to focus more on grow medias.
Edited by Ferather (12/02/18 07:35 AM)
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Ferather
Mycological



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Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather]
#25654762 - 12/03/18 02:26 PM (5 years, 2 months ago) |
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The next test I will make sure the wedge is not too close to the lid, sorry for the images. Regardless, I have lignicolous Cubensis on WL-Tek (cellulose) + tea.

Speed is improving over time.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25659712 - 12/05/18 04:21 PM (5 years, 2 months ago) |
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Growth is very slow, spending it's time and resources on slow release cellulose. This time it's not stalling like the 100% cellulose sample did.

Not as fast as WL-Tek + 8g sucrose.
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WL-Tek + Tea, +16 hours, with visible growth progression. T-Gel + MG-YN growth is weak. Evidently, the Cubensis sample grows better with cellulose than without.
Decay of the phenols appears to be partial, not complete. The sucrose in T-Gel test 1 improved growth.

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Too slow to be usable as spawn, and growth isn't as good as WL-Tek + sucrose. Cubensis does decay phenols as carbon, but it's not very ideal for it.
Edited by Ferather (12/08/18 10:40 AM)
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Ferather
Mycological



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Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather]
#25667298 - 12/09/18 07:52 AM (5 years, 2 months ago) |
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Growth on, T-Gel + MG + trace sucrose, is better than, T-Gel + MG + YN. It seems the additional sucrose makes up for partial decay of phenols.
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https://www.shroomery.org/forums/showflat.php/Number/25637557#25637557
Turns out it's not mold, and instead something else, it appears to have produced a sclerotium-tuber (rhizome).
It does not appear to be able to grow out, and seems totally cut off, so perhaps this is the reason. Regardless, it's not a mold as there are no spores, and no aerial stands with pods.
No idea what it is, but it's a mycelium, and it germinates on T-Gel.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25695235 - 12/22/18 05:53 AM (5 years, 2 months ago) |
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T-Gel recipe: 100g Water, 2.4g Agar, 1.6g Black Tea (0.6g Soluble), 0.8g Sucrose, 0.04g MG.
6 Weeks, I appear to have about 24 knots forming, here are a few of them.

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https://www.shroomery.org/forums/showflat.php/Number/24012223#24012223
Typically mycelium like around 25-30:1 carbon-to-nitrogen, in various formats.
Edited by Ferather (12/22/18 04:01 PM)
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather] 1
#25726604 - 01/06/19 07:01 AM (5 years, 1 month ago) |
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Exports from another thread:
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Decided to see if I can double the amount of tea solubles, note: increasing the amount of tea increases total acidity. Using my digital pH pen, I measured my test recipe, and measured the amount of CaCO3 needed.
Water: Hard water, tap, contains calcium bicarbonate, pH 7.2-7.5.
100g Hard water, 3.2g Black tea, 0.8g CaCO3 - pH 5.9-6.0. 100g Hard water, 3.2g Black tea - pH 5.5-5.6.
Note: The above result will vary (water, tea).
The increased solubles should be equal to 50% of the normal amount of ME used (1.5g instead of 3g). Currently T-Gel is roughly equal to 25% as malt extract, hence the optional flour additive.
Note: pH 5.5 should be fine for some mycelium, others may prefer higher.
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So I tried double concentrate with added CaCO3, but it did not set properly (precipitate), so I'm trying without CaCO3. Recipe: 150g Hard water, 4g Agar, 3.2g Black tea. Water + agar, first, then the tea bag is added.
I microwaved the solution using 25 second intervals, until 50g of water was lost.

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Standard 18-24 hours response
The blue tint generated by the added MG (wedge) is nearly gone, the wedge also looks fairly clear with penetrative light.

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The Cubensis is doing much better than I expected, almost as fast as T-Gel + MG. The double concentrate seems better. Growth is fairly thick (populated), and visibly penetrative, it's thickest on the new agar (at the moment).
The wedge surface (bottom) has signs of white-rot style decay, turning yellow (bleached). In addition, the wedge and agar have not turned dark like in previous tests.

No blue tint (wedge, MG).
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05/01/2019 - Similar speed to full strength malt extract.

05/01/2019 - Evening, plus 8 hours from before.

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Adaptation to pH and media composition, growth is no longer uniform, and now selective. The adapted growth is equal in speed and population to white-rot oyster.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25728574 - 01/07/19 07:09 AM (5 years, 1 month ago) |
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The adaptation has now passed around the wedge, which took a few days. All growth has now continued. Growth is populated (branched), not rhizomorphic (clumped), its mostly flat (not aerial).
Speed is a little slower than ME, however there's no weak growth.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25731157 - 01/08/19 09:12 AM (5 years, 1 month ago) |
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Today's images are a little misty due to a small circle of condensate directly above the wedge and growth, which indicates heat production. Some of the mycelium looks coloured by my torch and the agar, ignore that, all growth is pure white and not discoloured.
As mentioned yesterday all growth has continued, overall speed is average, faster than cyans on ME.
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Humanhelper
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Re: Cubensis [Data Log] [Re: Ferather] 1
#25731171 - 01/08/19 09:20 AM (5 years, 1 month ago) |
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Ferather, I completely dig what you are doing here. It is always good to get the brain chugging on this info. Enzymes are catalysts that lower the activation barrier of already favorable rxns. You mention pH which is important but it is also good to narrow down the temp range as well. These runs will proceed without but it is obvious the enzymes speed up the Ron’s considerably and could be favorable in certain recipes to isolate specific pH and temp that the specimen can also withstand. It would be awesome to see your hypothesis and also some temps since you’ve mentioned pH. I’m hoping to follow this closely and I may be able to assist with interpolating the data into a useable ‘recipe’. Keep up the strong work! HH
-------------------- It looks a lot more like it does now than it did...
 
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Ferather
Mycological



Registered: 03/19/15
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That's true I have not really covered temperatures with enzymes, mostly because I need regulating equipment to get a set constant temperature. Adding X to Y and then measuring the pH is much easier and cheaper than running equipment to regulate temps in my case.
I just run with room temperature (22°C) for everything, unless fruiting needs a different temperature.
Edit: I would probably also need to isolate enzymes and test them separately.
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Ferather
Mycological



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Posts: 6,325
Loc: United Kingdom
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Re: Cubensis [Data Log] [Re: Ferather]
#25733667 - 01/09/19 10:50 AM (5 years, 1 month ago) |
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Today's images, I recharged my torch (see here), and used a holder to produce a good angle.

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I conclude my Cubensis test.
Cubensis can and will target phenols-other, and decay them as the sole carbon source (no sugar, starch or cellulose). However to do so, it must also have the essential macro-micro nutrients present, with the carbon source.
The Cubensis produced it's own carbon materials from the phenols: glucose, proteins, so on.
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Notes:
Double concentrate required no additional nutrients, however it's slower than without. Sucrose, MG and YN had beneficial impact to the available pool of nutrients. The addition of nutrients improved growth and supported adaptation.
Edited by Ferather (01/10/19 02:09 PM)
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Ferather
Mycological



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Posts: 6,325
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Re: Cubensis [Data Log] [Re: Ferather]
#25739765 - 01/12/19 09:17 AM (5 years, 1 month ago) |
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[Recipe A]: 100g Water, 2.4g Agar, 1.6g Black Tea (0.6g Soluble), 0.8g Sucrose, 0.04g MG. [Recipe B]: 100g Water, 4g Agar, 3.2g Black Tea (1.2g Soluble). I used hard water *.


Some agar is lost to the removed tea and bag. * 150g microwaved down to 100g.
Edited by Ferather (01/12/19 09:30 AM)
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25745816 - 01/15/19 03:28 PM (5 years, 1 month ago) |
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The new plate (recipe B, above), has turned radial, growth is thick and populated, it's clean but not quick.

The master plate (recipe A, above), has bleaching or whitening, I removed the pins.
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Ferather
Mycological



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Re: Cubensis [Data Log] [Re: Ferather]
#25763438 - 01/23/19 02:43 PM (5 years, 1 month ago) |
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Cubensis experimental wedge to T-Gel (updated recipe, today). See my signature for the updated recipe. I have reverted back to Intralabs food grade agar. It's 250% better, and needs less.
I'm using T-Gel plain, which means no optional malt extract.
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