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senescence experiments with cubensis?


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#1 newmoon

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Posted 26 July 2021 - 01:50 PM

Hello brains,

 

The guideline I've seen is that cultures will eventually senesce and lose vigor after repeated transfers (a2a, g2g, clones of clones) with the same media. I've been searching on here to see if I can find some experiments showing how fast this happens, but am only seeing general guidelines without hard data.

 

I'm sorry if I'm missing it, but could someone point me to a previous experiment with cubensis looking at this? If not, I'm thinking I might do this and write it up in the myco lab.


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#2 newmoon

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Posted 28 July 2021 - 08:44 PM

Nothing so far, I guess? I'm thinking I'll do this, then, and use this thread as an in-progress workspace.

 

I've thought of three experiments:

1. agar -> grain -> substrate -> fruits -> clone to agar, repeat

keeping track of yields and colonization time for each cycle;

 

2. agar -> agar, repeat

fruiting every third (or fourth, or whatever) transfer, keeping track of yields and colonization times;

 

3. grain -> grain, repeat

fruiting each transfer (or every other or whatever) and recording data.

 

My thought is to do use three clones each of three cubensis varieties simultaneously (9 cultures in total). Due to space considerations, I think I'll fruit in vitro in bags. I prefer to grow on manure and straw generally, but to make things more consistent I might just use coir for substrate, not sure about this still.

 

The main thing I can't control is temperature: in the winter I grow in a heated space, so I can control that, but it gets hot in the summer and I don't have cooling available.

 

I have space to do one of these experiments at a time, currently. I'm curious about the results, but I also want to get a start at doing statistically rigorous experiments with psychedelic agronomy as there might be somewhat of a shortage of this in the OMC. I'd really appreciate feedback, suggestions, or critique...


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#3 newmoon

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Posted 10 August 2021 - 08:13 PM

OK, I'm going to do this. I'll start by trying to quantify how quickly cultures degrade after repeated agar -> agar transfers with the same media. Thinking I'll do two replicates each of three varieties, as that's about as much space as I can put towards this.

 

Swiped cubensis PESH, Costa Rica, and PE#6 spores to agar. I've grown PESH and Costa Rica before (the first for about a year now, the second for several years), PE#6 is new to me. I'm going to grow tubs without isolating strains, and then I'll pick a couple fruits to clone of each to get started.

 

agar.jpg


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#4 Sidestreet

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Posted 11 August 2021 - 06:06 PM

Science!  A well-documented thread with lots of good pictures would be a big deal.  :)


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#5 newmoon

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Posted 16 August 2021 - 11:56 AM

Everything germinated, and I did a transfer to clean it up. If those are clean, I'll go to LC -> grain -> tubs and pick out some clones to use.


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#6 rockyfungus

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Posted 16 August 2021 - 07:59 PM

I feel it’s only documented in the gourmet industry as they don’t mess with spores.

Who knows how old those numbered cultures are. I’ve transferred plates out for a year with no issues. I stopped marking after about T25 lol


Edited by rockyfungus, 16 August 2021 - 08:00 PM.

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#7 newmoon

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Posted 09 October 2021 - 10:30 AM

Had some things come up, but I'm finally getting going on this. I'm doing an agar -> agar experiment and a clone of clones experiment at the same time.

 

Here's the protocol I've settled on:

 

Agar -> agar:

Make a transfer every two weeks, grow out every other transfer, document yields and time from spawning to first fruit. Do this for two clones each of three varieties.

 

Clones of clones:

Take a clone of a fruit to agar, go to grain, spawn, fruit, take a clone of the fruit to agar, repeat. Do this for one clone each of three varieties, documenting yields and time from spawning to first fruit.

 

 

For both I'll be going with cubensis Costa Rica, PESH, and PE6. Agar is 0.3g LME, 0.3g agar agar, 15mL water in a 4oz mason jar, sterilized for 20 minutes @ 15psi. The grows will be agar to grain, which is 1 cup of oats and 2/3 cups of water in a quart sized mason jar, sterilized for 60 minutes @ 15psi. Substrate is equal parts straw, manure, and wood shavings, pasteurized at 145F for 6 hours. Each grow will be 1 jar of spawn and 1 full quart of substrate in a mycobag, fruited in the bag (given my space restrictions this seems the most feasible way).

 

I might do multiple bags of each grow, depending on space.

 

 

Here are the clones I selected for PE6; two for the agar -> agar experiment and one for the clones of clones experiment. Don't have the PESH and Costa Rica clones yet.

 

1.jpg 2.jpg


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#8 MushLuvR

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Posted 09 October 2021 - 06:43 PM

This looks like a Fun Guy project to me...I look forward to seeing your results @NewMoon... Awesome Start and All the Best Broseph.   :thumbs_up:


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#9 Sidestreet

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Posted 10 October 2021 - 09:32 AM

I'm really looking forward to seeing what you come up with, too.


Edited by Sidestreet, 10 October 2021 - 09:33 AM.

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#10 newmoon

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Posted 11 October 2021 - 12:33 PM

Thanks, y'all! I'm expecting this to be a pretty long project, but if I'm just doing bags I should be able to keep it going without it taking up too much space. Those clones have started crawling onto the culture media, and so I'll make my first transfers on the 22nd.



#11 newmoon

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Posted 12 December 2021 - 03:19 PM

Been making transfers every two weeks. Starting fruits for clones of PESH and Costa Rica are pictured.

cr.jpg pesh1.jpg pesh2.jpg

 

I've just started harvesting the bags from PE#6 transfer #1; here's one such bag. I'm finding that the first flush per bag seems to be 100-150g fresh (2/3 quart oat spawn, 1 quart substrate).

bag1.jpg

 

Is it a little slow around here recently? I've been very busy lately and haven't had as much time to stop by.


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#12 invisibilitysyndrome

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Posted 18 December 2021 - 12:47 PM

Good luck I do believe itll take a while for you to start to notice what your looking for.
In my exp things got better over time befor they got worse starting from multi spore but its a neat project.
What are the traits that you are going to be lookin after as proof?

#13 newmoon

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Posted 21 August 2022 - 11:44 AM

Guys I'm really sorry to have an anti-climactic end to this, but a series of illnesses and personal situations over the last year has left me unable to follow through with this in a rigorous way. I can say that there isn't obvious sign of degradation after growing out 10 transfers on the same media (MEA) over the course of 10 months; it would be great if someone else wanted to try this, but I think I have to give up. I guess from this very limited attempt I'm not going to be overly concerned about switching media or limiting transfers; maybe it's a problem at some point, but it doesn't seem to be an issue at the scale that would affect me (or possibly many of us).


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