Smellyhobbit said:

My plates have been acting very strange lately...Any ideas?

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Have you tried playing classical music for them?

Kidding. I got nothing.

Pic VAT: x7x+ plin looking uncircumcised, and budding a forest of plins.

MycCycle said:

CS works on similar principles but adds another element: the mycelium has to physically force its way through clean agar, which acts like a mechanical filter.
That tunneling not only blocks bacteria, but it also (supposedly) naturally selects for stronger, more aggressive mycelium.

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2024-12-11 said:

Thanks!! That is a very helpful write up. I love CS for my own agar work already after running a handful of cultures through the process, and seeing the results others are having here. You’ve expanded my understanding of it a good deal here too.


I’m still very curious about CS without a roof specifically.

MycCycle said:

CS works on similar principles but adds another element: the mycelium has to physically force its way through clean agar, which acts like a mechanical filter.
That tunneling not only blocks bacteria, but it also (supposedly) naturally selects for stronger, more aggressive mycelium.

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The roof ensures the mycelium has to tunnel to get out, ya? So doing this without a roof, the mycelium does have to cross the gap on a new plate (better than just trenching on the old plate) but it can crawl up the side of the agar without needing to tunnel, ya? Sorry if I’m being pedantic. If there is a reason to omit the roof I’m all ears but at that point it isn’t really CS, right? CS benefits from tunneling but CS without a roof doesn’t force tunneling so is it CS?

Sorry if I come across as a bother, I am trying to learn and I appreciate your earlier response very much.

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No bother at all, these exact same questions made me question the process.

I had an issue with accepting the fact that Grzybowski presented this process with an improvised agar "coverslip" instead of an actual coverslip, which was never intended or tried by the authors of the research paper, as far as I understood. It went to a point where I reached out them and asked for clarification. They confirmed that they used a glass coverslip.

The glass coverslip's purpose is to isolate the tissue and force the mycelium to grow outwards.

A byproduct of this is the tunnelling, which may/may not act as a mechanical filter.
A byproduct of using an agar coverslip, is that you give the bacteria another medium to grown on, and having no glass coverslips, I decided to skip this step.
I'd like to mention that all three variants seem to work. I prefer using the roofless cabin, as it's the least work and satisfies my needs.


I'm no biologist and have no academic competence to back this up, but my understanding of the concept, in short:

Bacteria can latch onto mycelium to help with its motility and expansion even though it needs biofilm/moisture and a medium that is soft enough to move in.
Bacteria cannot stretch through air and open spaces, unless it's somehow blown there or moved with moisture.
Creating a gap like that itself should therefore be sufficient to shake off any potential bacteria as bacteria don't really have any mechanics that allow for it to latch on to the mycelium through the gap, and as you can see on the x7x(+) plate, it almost looks as if one of the sides decided that "Alright boys, we focus on this direction", which to me is a sign that there is something more aggressive there, than throughout the rest.


As mentioned earlier; I've found my success rates increase, times and resources spent shortened after doing this throughout every transfer.
I am only stating correlation here, not causation. As it takes a lot more work than what I have put in, to state as a fact that the sequestering is the only/main reason for the improvement.


I took these now as an illustration, so it's Ochra, Bluey and x7x(+), to show the different stages.

AZ Starman2112 said:

So some of you might remember I posted a couple pics of a plate that had bacteria on it and was being taken over by aggressive rhizo growth from ochra/nats. Well I never tossed that plate because I was just curious to see what would happen to it. That aggressive myc eventually took over the entire plate completely covering and masking the contam. I left the plate to be and still refused to toss it, just for experimental experience. Well I was away for four days and came home to this last night… It now has a bunch of plins on it. What would you do? Do I open it and transfer the plins to new plates giving them new life? Or just let them go on and live out their life in the plate along with the contam hiding underneath the myc? Check out the following photos…

This was the plate back on April 24th which was inoculated on 3/28 with a spore print…





This is the plate tonight on May 14th…

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In addition I found this this morning on a plate that I took transfers from a couple weeks ago. I put the plate at the bottom of the pile figuring I’d eventually just toss it. The T1 plates have gone aggressively rhizo, and ready to go to grain, and then I saw this! Wondering now what to do with the plate I referenced above as well as this Mazatapec giant plin?

Huskies said:

I know all of our shared scientific knowledge says it should fail, but my inner wizard trusts the mycelium.

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MycCycle said:

Does any of you use or find the 2 and 3 compartment plates useful?

I know RR used these in some of his videos, but I can't remember having seen anyone use them here, since I started lurking

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Maybe, I do sometimes put more than one transfer on a plate.


Bispo


Or maybe for something similar to cabin sequesteration, make it climb over a wall instead of out of a hole. Dunno, just spit balling.
I'd probably find excuses to use them if a had them, but don't think I'd go out of my way to stock them.

MycCycle said:

Does any of you use or find the 2 and 3 compartment plates useful?

I know RR used these in some of his videos, but I can't remember having seen anyone use them here, since I started lurking

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You don’t need compartments to run more than one sample on a plate. You just have to move them before they run into each other. I like to do this when I take multiple samples from spore inoculation. It lets me see what each will do, and move the good ones to their own plate.