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The Master Plan (A quasi-meta-tek) * 11
    #28473547 -

Preamble


The 5 objectives, as I see them:

1. The ability to start, clean up, test, compare, expand, and preserve culture lines.

2. The ability to complete the entire life cycle multiple times in a short period.

3. The ability to simultaneously harvest fruit from a wide array of different cultures. Either to test them for potency concurrently, blend them together for a stable average, or a combination of the two.

4. The ability to maximize use of available containers and space without creating bottlenecks or impossible work situations.

5. The ability to maintain all of the above without constant deliberation, and balance it against having a life, leaving town, etc.

Underpinning this is the belief that repetition is the best teacher, and that the best way to deal with a failed project is to start another one.

In this thread, I'll be posting some scheduling ideas that I geeked out on hard this summer.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28473562 -

Will be looking forward to this one :popcorn:

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Re: The Master Plan (A quasi-meta-tek) [Re: Womble]
    #28473611 -

Quick note on parameters

My current general technique:

Agar work until I have something worth dropping onto grain (agar work will be discussed later)

Dropping agar wedges into small grain masters

Expanding those grain masters into quart jars

Running g2g transfers with my quart jars, 1 to 3 new jars from each old one

Using the remaining grain from the donor jars for spawning to shoeboxes


As this is what I'm currently doing, the language I use will be based on these practices. However, in the abstract sense, these ideas are applicable to any container or setup. I will also be quoting times based on my current experience with my cultures in these vessels. Those, too, may vary depending on the particular situation.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven] * 1
    #28473657 -

An approach to cultures and g2g transfers

The only actual technique here which might be significantly different from the norm is the way I handle g2g transfers.

Cultures can easily be expanded while on agar plates. And one plate can provide enough wedges for many master jars. One small master jar can easily inoculate three or four quart jars. And I'm typically trying to run multiple cultures. So, from my point of view, there's no point in using g2g transfers to expand a culture. I've had ample time to expand it by the time I get to my G1 jars.

Instead, I just use them to perpetuate it, i.e., one transfer from the jar and then I go on to spawn it.

Any culture theoretically has the potential to expand into thousands of jars. The limitations are senescence, contamination, and our own willingness and ability to do the expansion work. We all have good days and bad days, hit the lottery one day and the red light camera the next. We need time to run things out and figure out who our rockstars are. We need to keep our eyes open and watch for contamination creeping into our lines. And, when it comes to g2g, we need to watch for signs that the old grain hanging around in our later transfer jars is starting to throw the whole thing off and let the nasties in.

So, my approach is predicated on two thoughts.

The first is rather simple. It's better to use a small amount of grain for a transfer, because over time that means you'll have fewer old grains hanging out in your jars. Also, it'll make it easier to shake the jars, and easier to monitor your inoculation points after that first shake. So, then, why not use your spawn jars as g2g donors? The amount you should use to inoculate a new jar won't really be missed at spawning.

The second is turning the concept of growing around. We know we want to grow cultures out. We know we won't stop until we reach our goals. We know that, once we reach our goals, we'll just find new goals to obsess over. And we know that any one good culture has an expansion capacity that far exceeds our ability to do the work. So, instead of looking at cultures that we may expand if they seem promising, I think in terms of basic growing units. I don't have a dozen cultures I want to grow out. I have a pool of agar hanging out at the job fair, and maybe 10 masters I want to fill. If the field is tight, that may mean 10 different cultures from 10 different plates. If most of my plates are dogshit, that might mean two different cultures from three different plates. Etc, etc. And that principle applies all the way down through later transfers. When it comes to perpetuating quart jars, this becomes especially useful. Let's say I got 10 quarts lined up for a round of g2g transfers. Then I take a closer look at them and realize that 2 are contaminated, and 4 are from culture lines I don't care to continue- ok for spawning, but that's it. Even though I'm down 6 donor jars, the 10 jars I just pulled from my Presto won't go to waste. All I have to do is pull a few extra transfers from the 4 remaining donor jars, because, as I mentioned earlier, I can pull up to three transfers from a myco-quart and still not really notice the difference at spawning.

In other words, I can develop an expansion and production schedule that's independent of the cultures I actually have on-hand, and is scaleable on-the-spot to account for changes in circumstances. This also allows me to run cultures in different stages of development at the same time, and be ruthless about eliminating lines that are showing red flags.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28473679 -

Basic numbers

You need containers, and then you need a place to put them.

There are two absolute limits:

-The total number of quart jars and lids (in my case, lids are the limiting factor, I'm very picky about them)

-The total number of tubs

While there's always room for improvisation and expansion, I think it's important to consider and determine these two limits, and treat them as fairly absolute. When you're drowning in spawn, it's a lot harder to un-ring that bell.

There are also soft limits worth considering:

-The total number of jars you'd be ok with spawning in a given week

-The total number of tubs you'd be ok with actively harvesting in a given week

-The total number of PC runs and g2g transfers you want to commit to- weekly, monthly, and annually

Each vessel takes time to fully colonize. At which point it's ready to take transfers from, spawn, or harvest.

Small grain masters
-time to full colonization: 3-4 weeks from drop
-optimum transfer window: 4-6 weeks from drop

Quart jars
-time to full colonization: 3 weeks
-optimum transfer window: 4-5 weeks
-optimum spawn window: 4-6 weeks

Shoeboxes
-time to full colonization: 2 weeks
-main harvest window: 3-4 weeks
-ready for transfer to secondary fruiting chamber: 3 weeks
-time to discard: 4-6 weeks

All of the above numbers provide a foundation for construction of a master calendar.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28473771 -

Refined numbers

Ideal g2g interval: 30 days
Ideal spawn interval: 35 days
Expiration point: 60 days
Blackout period for spawning before extended travel: 35 days


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven] * 1
    #28473864 -

Most basic example

Let's say we have an agar plate we want to grow out, representing one culture line. Furthermore, for the sake of simplicity, let's say that we won't bother with a smaller grain master, and we're dropping straight into a single quart jar. And let's assume that we only have 2 quart jars and 1 shoebox to work with.

In this case, we'll say that we want as much of a continuous process as possible, and that we're willing to extend our culture out to 4 generations of spawn.

Day 1: drop agar wedge to quart

Day whatever, whatever, and whatever: shake and monitor quart jar, to speed colonization and watch for issues

Day 28: Soak grain (if that's what you're into)
Day 29: PC
Day 30: Transfer from colonized jar, creating 1 new jar. At this point, we're at our peak quart capacity, since our limit was set at 2 jars.
Days 32/33/34: Do whatever is needed to prepare for spawning (coir prep, tub/space prep, etc.)
Day 35: Spawn our original quart jar. We are now at peak tub capacity, as that was set at 1. But we've freed up a quart for the next round.
Day 50: First shoebox colonized.
Day 55-65 ish: Peak harvest from first shoebox
Day 60: Transfer from colonized jar ( back at full 2-quart capacity, with one quart ready to spawn)
Day 65-70: Point where the first shoebox should be available, and the next quart can be spawned.
Day 80-85: Second shoebox colonized.
Day 85-100: Peak harvest from second shoebox
Day 90: Transfer from colonized jar (3rd quart to 4th quart). Back at peak quart capacity.
Day 100-105: Point where the second shoebox should be done, and the space available for another round
Day 120-125: Third shoebox colonized
Day 120-135: Peak harvest for third shoebox
Day 135-140: Point where the third shoebox should be done, and the space available for the fourth and final round
Day 155-160: Full colonization for fourth shoebox
Day 155-170: Peak harvest for fourth shoebox
Day 170-175: Fourth shoebox should be done

So, according to this simplistic and idealized calendar, we've done four tubs of the same culture spread out over a 6 month period. We've had ample opportunity to pull prints and clones, run bioassays, and also watch our culture expand into grain jars while simultaneously seeing how it performs in tubs. If we're slacking on the agar preservation side, then we've also had ample opportunity to pull grains from our jars to put back on agar. We never went above our absurdly low limit of 2 quarts and one shoebox, and we never had to spawn or harvest more than one tub at any given time.

While the last couple jars had to sit around for a little longer before spawning, they were nowhere near our (arbitrary) expiration date of 60 days.

Due to the limitation of only having one tub, we had to work around the vagaries of exactly how long any given shoebox might last. But even with that, we've only seen a modest amount of drift into uncertainty, and it still hasn't gotten to the point where it might impact the availability of quart jars for the next round. By adding a second tub, providing a secondary vessel to transfer the old substrate into, or simply insisting on a definite expiration date for old tubs, we could eliminate that wiggle room. And make even the spawning schedule set in stone.

Of course, by only doing single sequential transfers from one plate, we've left a lot of dead space in the calendar. We've also created a situation where, if anything goes wrong, our one and only line is dead.

But of course, almost nobody would consider running g2g transfers one quart at a time. And we probably have multiple lines we want to run. If we assume, say, a run size of 10 quarts in the Presto, then we could increase our quart limit to 20, and our tub limit to 10. The same schedule would apply, but we'd have the ability to fill in any discontinued lines by pulling more transfers from the remaining ones.

And if we add another set of lines that runs on a similar but offset schedule, then we can create a situation where something is happening almost every week.

The key metric to keep track of in planning a more complex calendar is the total quart count, and the total tub count. Your peak capacity will be the ultimate limitation, followed of course by the amount of work you're creating for yourself.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28474170 -

Want to spread the sterile work out, but truncate the spawning

In this example, we'll assume that we want to start putting some of our cultures on grain, but we've got an out-of-town trip on the horizon and don't want to get involved in spawning yet. We figure that we'll be ready to throw down on some spawning when we get back from Cave-In-Rock or wherever, but that's a ways off.

So, based on our numbers, we know that we wouldn't want to spawn anything once we're less than 35 days from our trip (helicopter rides!). And when it comes to making spawn jars, we'll need to keep that 60 day expiration date in mind.

Just to make things more realistic, we'll assume that we're running 10 quarts at a time. And to make them more complex, we'll assume an offset schedule involving two different sets of culture lines.

We'll keep the sourcing fuzzy, and just assume that we have a bunch of masters hanging around to inoculate the g1 of each line. And switch to a weekly format for clarity's sake:



So, with this 6 month calendar, we've spawned 60 shoeboxes in a 10 week period. We've given ourselves a ramp-up period to start two different chains of g2g work (1 and 2), and then run all our lines out to g3 (transfer c). If any of those lines crap out or look funky, we can simply use other lines to fill out the ranks. And we've carved out two weeks with zero responsibility.

Now, most critically, I've set week 1 as being about 8 weeks out from our initial spawning. If any more time elapsed, then our first round of jars would be past its (arbitrary) 60 day expiration date. But this way, we can be spawning representatives from all our lines in the same week, and then quickly be following up with their offspring.

There are still some blank spots in the calendar, and naturally things tapered off after we stopped doing any more g2g transfers. Of course, the cycle could be continued indefinitely for as long as we felt comfortable running out those transfers. And if we added a third set of lines to the mix, we could fill in more space. But that would naturally require a higher overall capacity, and even more care to avoid running into expiration dates.


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Edited by B Traven (09/18/23 07:46 PM)

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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28474532 -

:popcorn:


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Somewhere between chaos and disorder.

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Re: The Master Plan (A quasi-meta-tek) [Re: med man]
    #28474625 -

:threadmonitor:

I like your thoughts and timeline


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If you have used a Miraculix Psilocybin QTest, could you please share your results?

Shipping free Ps. natalensis spore prints to any address in The Netherlands, just :pm:

:mushroom2: Mush love :mushroom2:

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Re: The Master Plan (A quasi-meta-tek) [Re: Tiamo]
    #28475061 -

Running hard, but with some pacing

In this example, we're not trying to carve out any time off, and we're ok with a couple months of heightened workload. But we're still trying to spread out our spawning and transfers, keeping everything steady. This will give us ample opportunity to assess various culture lines, and fill in blanks when things screw up. I've run this out through g3 again, but the pattern could be repeated past that if we wanted to run our lines out further. Or, another option would be to introduce new ones from our stash of plates and masters.



The asterisks on this calendar represent points where we could reduce the total number of quart jars needed. In week 5, we make our first g2 transfer (for set 1). Once we've done that, we could dump those quarts into secondary containers, or even just spawn them early. From there through week 11, we could simply spawn before running our transfers for the week, which would also free jars up early. So it would be possible to get away with running this entire schedule with 50 quart jars on-hand.

The arrows represent points where there's more flexibility with the spawning calendar. After week 12, we no longer care about freeing up jars, so the only time limit we're working against is our (arbitrary) spawning deadline. We have until around week 22 before the last set of quarts expires. If we had a lower capacity than 50 shoeboxes (or the equivalent in monotubs), we could simply abandon the spawning schedule at that point, and refill tubs as they became available.

Here, we've managed to keep our g2g work steady at 10 quarts a week, while running 4 different sets of culture lines (40 total). They might represent just a handful of lines expanded out from a few masters, and in that case, assuming all goes well, a lot of people might be looking to spawn multiple quarts to a larger tub. Or they might represent someone's attempt to test and grow out 40 different lines at once. Either way, we've run 120 tubs, and learned a lot about the cultures we're working with. Just about anyone who could commit to this work schedule could move through it, and hopefully find the experience rewarding on many levels. But of course, the main determining factor would be how robust their starting cultures were.

Which brings us back to agar and masters.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28475797 -

:thisisgonnabegood:

I m hooked !

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Re: The Master Plan (A quasi-meta-tek) [Re: Traveled] * 1
    #28477673 -

Agar

I see agar work as a microcosm of grain and spawn work. You can manipulate, select, and expand cultures in a tiny space. In the span of a couple months, you can create a lifetime of potential work. It provides a low-stakes way to play around and look for something good. And it contains infinite possibilities for expansion. A single plate of germinated spores could lead to dozens of distinct clone lines. Combined with the expansion possibilities we've already discussed, that could easily represent thousands of jars.

So, yeah.

It's fun, and the more you do it, the better you get at identifying clean cultures that are ready to move to grain. Also, you can use cold storage, slants, etc., to preserve agar lines you particularly like.

Also, our plates are the foundation for all future work. A real garbage-in, garbage out type situation. The stakes are low when playing around with agar, but they quickly escalate once we place our bets by moving it to grain. It seems prudent to have lots of back-up options, in case it turns out that we bet on the wrong teams.

And that typically all leads to obsessive hoarding, and a pile of plates that the cultivator is completely overwhelmed by.

I believe that it's ideal to pull germinated mycelium off of a starter plate as soon as you possibly can, to begin the culture differentiation process and get it away from any contams that might also be germinating.

For later transfers, I find it ideal to pull mycelium from a plate as soon as its leapt from the starting wedge and can be cut from the new agar. Well before it hits the edge of the container. Again, this is the best way to outrun contams.

So those both require one to be ready and alert. Next generation of plates free, a reasonable number of plates to monitor, etc.

And, of course, the obsessive hoarding situation is somewhat antithetical to that.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28477682 -

My personal solution is to employ the same container management philosophy that I do for grains. I'm not running different agar lines, I'm managing a certain number of plates.

I do no-pour agar in an instant pot. Boil up a batch of agar, load it into the instant pot, and go to bed. The next day, (or the day after that if things get away from me), I can pop it open and run my transfers. I find that convenient, but also, the fact that I can't fit very many plates into my IP helps provide a limit and structure to my agar work.

Everyone has their own preferences when it comes to developing culture lines. Right now, I'm really into plate pins. I know some people hate them, and some people want to clone fruit after they've had a chance to see it grow. I personally like plate pins, and haven't had any really bad experiences with them. At this point, with the varieties I'm running right now, I'd rather grow out an untested plate pin than a multispore culture. I also find them very convenient. So for the purposes of this discussion, I will be using my current practices as the baseline. It should be fairly easy to imagine how one would shift things around if they were working from straight multispore plates, taking clones from their tubs, etc.


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Edited by B Traven (09/21/23 07:12 PM)

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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28477696 -

Basic numbers

Time to germination/transfer from a scraped plate: 1-2 weeks

Time from one agar transfer to the next: 3-5 days

Arbitrary minimum number of transfers from germination: 5

Arbitrary minimum number of transfers from a cold-stored plate: 3

Arbitrary minimum number of transfers from a cloned plate-pin: 3

Time to full colonization and formation of plate pins on a nice clean plate, if all goes well: 3-4 weeks

My logic behind a minimum of 3 transfers is this: The first transfer still has the original material in it. The second transfer has material that was in the same room as the original material. The third transfer should be good to go, unless there's some bionic contaminants present. And, mind you, I'm talking about transfers from a clone plate or a stored plate, i.e., a situation where we already have the genetics we want, and already did our initial clean-up transfers long ago.

These numbers are then the building blocks for scheduling transfers. The temptation to expand cultures can be enormous, and scheduling plus a limited number of plates provides a way to keep that in check while meeting our goals.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven] * 1
    #28477716 -

Expansion

So, the core of agar work, to me, is running sequential transfers to get clean and organized growth. That is naturally a somewhat open-ended process, as there's no simple universal answer for when a plate will look good enough to move forward with. If we think back to the original example of one single g2g line, the same principle applies here. If we only take one transfer from a plate, then dedicating two plates to a given culture line is enough to keep transferring it indefinitely. We could even take that to the point of saying that when we drop agar from that line onto grain, the final plate could be put in cold storage and the empty one could be set aside for whenever we revive it.

So, that's pretty cool. 2 plates could cover an entire line, indefinitely. But there are two issues there: first, those two plates would then be tied up, indefinitely. And second, if anything goes wrong, then we've lost the line.

So there's one major reason we want to expand culture lines- to have backups. Others might include giving them away, creating a foundation for multiple g2g lines, or just creating a larger pool for potential plate-pins. There's also the likelihood that multispore plates are continuing to have their genetics narrowed by successive transfers. So in that case, expansion is a way to play around with more of the genetic soup on a given plate, and create bifurcating culture lines.

But every time we expand, i.e., take more than one transfer from a plate, we're creating a situation where that new line will either now occupy two plates indefinitely, or eventually be discarded. Every time we think "Oh, well, I'll just throw one of them in the fridge," we've taken another plate out of circulation.

So in my opinion, it's a good idea to think about exactly when and why we'll expand our lines.


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Edited by B Traven (09/21/23 02:18 PM)

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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28479698 -

Random example

Let's imagine that we're starting from scratch with spores, 3 to 4 months before launching one of the previously covered g2g schedules. The following schedule provides a rough outline of our agar work, with the following assumptions:

-A run size of 8 plates for our no-pours
-All plates are discarded after transfers
-Plates used for grain drops are placed in cold storage after the wedges are taken
-Each of the plates we select for grain drops will be used to inoculate 2 masters



You can see here that we've got two primary expansion points: when doing our initial few rounds of transfers after germination, and the initial transfer after we've taken clones.

There are several points worked into the calendar where we will be selecting our best-looking plates, and discarding others. Most notably, weeks 7/8 and weeks 15/16.

We also have a slight expansion in week 5, which could involve a variety of permutations. But most likely, it would involve discarding some plates and expanding others.

Of course, we could end up naturally eliminating plates from further work at any point in the process, as contamination or finky growth turn up. And the exact timing of our transfers and drops will likely end up moving around a bit. Agar work will be the leadt predictable part of the process, and any calendars we develop will necessarily end up being just guidelines. But the most significant part is making sure we're not drowing in containers, and have both the time and space to do all our work promptly.

It should be noted, once again, that this example assumes we're starting from scratch. In the case where we have some cultures already hanging out in cold storage, we could greatly reduce the time needed to get to our first grain drops.

This example also assumes an idealized situation where the number of minimum transfers I initially quoted ends up being enough. If more transfers were needed to get to send-able growth, then the process would be extended. However, based on the pattern of the work, the peak number of plates needed shouldn't change. On the other hand, one could modulate that a bit by adopting a ruthless attitude towards their plates, eliminating the ones that need more work, and filling in with the better/cleaner ones.


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Edited by B Traven (09/23/23 10:17 AM)

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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven] * 1
    #28479751 -

Masters

In the previous example, we'd have multispore masters ready about 12 weeks in, and clone masters ready about 20 weeks in.

Once we have fully colonized masters available, we're ready to begin a g2g schedule. They can also easily be included at any point when we're doing g2g work, if needed/desired.

Masters represent the point where the rubber meets the road. We've picked some cultures to bet on, and now we're starting our g2g journeys. Small masters (say, 1/2 pint or so) don't involve very much grain, and they offer us an opportunity to watch how our cultures behave on a new medium. Now is when we're watching for telltale signs of bacterial infections we might have missed, among other things. Masters can be kept in cold storage, and also just tend to have a slightly longer window of viability at room temperature. Also, you can always take any extra duplicate masters you have and spawn them to small containers, which also gives you an opportunity to test your cultures in fruiting conditions before you've invested much in them. So there are a number of reasons to hedge your bets by cranking out extra masters. But I think the best one, for me personally, is just having lots of options, so I can be absolutely ruthless about only using the best-looking specimens.

I won't write up a calendar for grain drops, but of course we should still be mindful of how many containers we have available for them, and roughly how many we want to be playing around with. This is perhaps most important for figuring out when we might be going overboard with the agar work, and creating plates that have no place to go.


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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28479764 -

Overlapping

One final note:

The agar work I outlined, while including many small runs, really wouldn't be all that heavy. In any given week where one is doing g2g transfers, adding a few agar transfers and/or grain drops to the sterile work shouldn't be a very big deal.

So, there are many different ways that one could work up a schedule that involves continuous agar work alongside the g2g's and spawning.

My personal preference is to layer everything in together,and then perpetually select for the best-looking vessels/lines. If a G3 jar is looking cleaner than the G1 I just made from a new culture, then that new G1 jar might be tossed, or relegated to spawning with no transfer, while the solid-looking G3 goes on to G4.


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Edited by B Traven (09/23/23 10:51 AM)

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Re: The Master Plan (A quasi-meta-tek) [Re: B Traven]
    #28479773 -

Alright, I think I hit all the major points I wanted to. I have plenty more to say, but at this point I'd be likely to start rambling and/or excessively repeating myself.

Any questions or requests for clarification are welcome, I'll do the best I can.

It would also be dope to see this thread turn into a general discussion about scheduling. Or for there to be another thread about it. I'd love to see how some other people deal with this stuff!


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