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PrimalSoup
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Registered: 11/17/09
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recommended # trial isolates for trait selection?
#21948857 - 07/15/15 06:42 PM (8 years, 6 months ago) |
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Just wondering if people have a preference when testing isolates for traits. My plans are to select out and isolate about 10 substrains from a PE swab I've saved in the fridge for a couple years (those are the original fruits in my sig). The swab germinated contam free in around 5 days the last time I put it to agar. The genetic variance is what I'm looking to capture of course, in this case likely potency but more along the lines of just the kind of trip I like. 
An ancillary question is this - IME it takes a little while for dikaryotic myc to show up on the innoc plate, but without resorting to microscopy on the plates is there a "best time" to start isolating from germination? As soon as growth starts or as it begins to proliferate?
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
Edited by PrimalSoup (07/18/15 01:14 AM)
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Kizzle
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Re: recommended # trial isolates for trait selection [Re: PrimalSoup]
#21957009 - 07/17/15 05:05 PM (8 years, 6 months ago) |
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It doesn't really matter unless you're wanting to isolate the monokaryotic mycelia for breeding which should be done early on.
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PrimalSoup
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Re: recommended # trial isolates for trait selection [Re: Kizzle]
#21957192 - 07/17/15 05:44 PM (8 years, 6 months ago) |
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Thanks, that's what I thought.
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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maddchef
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Re: recommended # trial isolates for trait selection [Re: PrimalSoup]
#21958316 - 07/17/15 09:52 PM (8 years, 6 months ago) |
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A preference for what when testing isolates? How many to test at a time, best way to test, substrate, etc?
I don't try to keep up with more than 10 at a time. I make containers that require no maintenance per my sig, easier than cakes, and let the iso run through whatever sub I will eventually use to grow it regularly if it turns out to be a winner.
Testing potency takes a while since you really should have a single individual testing and have to account for serotonin levels and tolerance.
Or......are you simply asking what traits are desirable for an isolate?
-------------------- In the land of the blind, the one eyed man is king. All mushrooms are edible, but some only once..... Easier than cakes I do science and shit.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: maddchef]
#21958870 - 07/18/15 12:24 AM (8 years, 6 months ago) |
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"Recommended # trial isolates for trait selection?" just your preferred number to run not how to do it. 
I'm looking for a number that gets a good cross section of variability without going to far to extremes - somewhere between 10 to 100 I expect.
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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36fuckin5
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22004116 - 07/27/15 11:46 AM (8 years, 6 months ago) |
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If you want the best one, run them all.
Personally, I just do a MS run, take a clone then isolate from there. You'll have a lot less strains to work with, and they'll all usually be decent if not great.
-------------------- Redd Foxx said: If you're offended I don't give a shit and don't come see me no more. Pat The Bunny said: A punk rock song won't ever change the world, but I can tell you about a couple that changed me. bodhisatta said: i recommend common sense and figuring it out. These are the TEKs I use. They're all as cheap and easy as possible, just like your mom.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: 36fuckin5]
#22004169 - 07/27/15 12:02 PM (8 years, 6 months ago) |
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Well, since I always isolate based on the myc appearance on the plate, I'll take anything interesting and see how it grows there. But what I don't and haven't done is to give less spectacular myc a chance, but my experience over the years selecting vigorous growing sectors suggests that that's the best way to do it in any case. Tomentose cultures (that remain that way) rarely have done well for me - I have one now from a free PFC syringe that is just this side of a complete failure.
My clones usually contam so I'm not really into that.
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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36fuckin5
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22004255 - 07/27/15 12:21 PM (8 years, 6 months ago) |
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Well yeah, if you're gonna isolate from spores you'd obviously take the best-looking, most aggressive growth. And then I'd grow ALL of those out.
If your clones are all contaminating you're doing something wrong. They're about as likely to contam as starting from spores IME. I won't do less than 5 plates from tissue, then isolate from there.
-------------------- Redd Foxx said: If you're offended I don't give a shit and don't come see me no more. Pat The Bunny said: A punk rock song won't ever change the world, but I can tell you about a couple that changed me. bodhisatta said: i recommend common sense and figuring it out. These are the TEKs I use. They're all as cheap and easy as possible, just like your mom.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: 36fuckin5]
#22005985 - 07/27/15 06:19 PM (8 years, 6 months ago) |
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No I just don't work with clones much, my experience hasn't been good so I never bother. 
My spore plates germinate contam free most of the time, taking something out of the open air and extracting a contam free portion doesn't work for me.
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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Kizzle
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22007829 - 07/28/15 02:14 AM (8 years, 6 months ago) |
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Quote:
PrimalSoup said: No I just don't work with clones much, my experience hasn't been good so I never bother. 
My spore plates germinate contam free most of the time, taking something out of the open air and extracting a contam free portion doesn't work for me.
You can take the pins from a MS agar plate when it starts pinning so there's a higher chance of getting a sterile culture. Probably a more vigorous culture too in terms of senescence compared to cloning pins from a fruiting bulk substrate.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: Kizzle]
#22009387 - 07/28/15 12:31 PM (8 years, 6 months ago) |
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Yeah, I've been considering that. But is there an advantage to cloning pins (at the end of the growth cycle regardless) over just selecting rhyzo myc for vigor? Perhaps the speed to pinning?
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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Kizzle
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22009728 - 07/28/15 01:36 PM (8 years, 6 months ago) |
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So you can get a genetically homogeneous strain. It's only way get one from a MS culture with complete certainty. Even though MS cultures produce sectors of a sort, sectoring between genetic individuals only occurs between incompatible single spore monokaryotic colonies.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: Kizzle]
#22009773 - 07/28/15 01:44 PM (8 years, 6 months ago) |
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But any given fruit from MS is just as likely to be a mosaic, SFAIK, regardless of whether it occurs on a plate or in a fruiting sub. If you know that after this step the resulting culture exhibits all the traits of an isolate then I'm interested.
My experience has always been that repeated selection of a sectoring plate always results eventually in a homogeneous culture that does everything an isolate is supposed to do...
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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Kizzle
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22009975 - 07/28/15 02:26 PM (8 years, 6 months ago) |
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You usually don't need any extra transfers with a sterile clone. Here's an agar pin clone. It may not look like a monoculture because of the asymmetry, that's in part because it's adapting to a new growth medium, but given enough space it grows into a perfect circle. Not that a perfectly circular colony is necessary to ensure consistency between substrains of a culture but it's a sign that there are no hidden contaminants.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: Kizzle]
#22010112 - 07/28/15 02:49 PM (8 years, 6 months ago) |
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Not about contaminents IME - the even circular growth is a culture that doesn't sector because it can't. I generally transfer until I have regular growth - my trial experiment here (which I've never done, which is why I'm asking) is for PE isolation and testing. - but not necessarily until it's completely regular.
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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Kizzle
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22010500 - 07/28/15 04:01 PM (8 years, 6 months ago) |
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Actually I'm not even entirely sure that two dikaryotic strains can't grow together without causing any sectoring.
What I am sure about is despite what a lot of people on this forum believe, and I used to be one them, a mushroom is composed of a single strain. So there's no need to keep doing transfers with it until the sectoring stops to know you have a single strain like you would if you were trying to isolate a strain from the mycelium or to know that it's not contaminated like you would with a nonsterile clone.
Although it's possible to wind up with multiple strains from a clone if it is sporulating or if you clone tissue from the very base of the stem.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: Kizzle]
#22010872 - 07/28/15 05:19 PM (8 years, 6 months ago) |
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Quote:
Actually I'm not even entirely sure that two dikaryotic strains can't grow together without causing any sectoring.
I may have to give that triple negative some thought. But, I'll grant that if they were very similar it would be hard to pick them apart on a plate. Which brings me to the next step, I suppose, mating individual monokaryotes with the aid of magnification.
For the other though, I can sort of accept that, given the way pins form. But I'll have to succeed at cloning sometime to see it work. 
FWIW I had a similar sort of situation come up some years ago while attempting to ressurect a stored mutant Ps. cyan print (15 years in the fridge). After some soaking I got sparse germination, with what appeared to be no more than a few germinations and only a couple of rhyzomorphic growths. I transferred these to the same plate to see what would happen. Got a beautiful example of incompatible growth, all from the same parent print:
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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Kizzle
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup]
#22011523 - 07/28/15 07:11 PM (8 years, 6 months ago) |
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It also seems kind of self-defeating to try and attain a vigorous strain by using a method that requires a crapload of transfers. Each transfer brings you one step closer to strain degradation. A couple inches of growth may not seem like much but it's about the distance the mycelium has colonize after a g2g.
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PrimalSoup
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Re: recommended # trial isolates for trait selection? [Re: Kizzle]
#22012444 - 07/28/15 10:11 PM (8 years, 6 months ago) |
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The transfers take only short periods of growth IME to show sectoring, less than you'd let a plate grow out. I finally have a use for my glass petris again 'cause I'll be making a lot of plates serially to process this stuff. 
I haven't done it but forcing a pin to go back to vegetative growth would seem to exact a toll on vigor as well. But senescence (if your talking about that) doesn't ever seem much of a factor in plating. Again that's just my experience.
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if you stand too close to the machine it'll start to eat youPrimal's simple tested teks and projects: Wheat Prep 2.0 Acidic Tea Tek Potency Project!
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36fuckin5
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Re: recommended # trial isolates for trait selection? [Re: PrimalSoup] 1
#22012512 - 07/28/15 10:32 PM (8 years, 6 months ago) |
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CLones definitely sector and are composed of different strains. Isolate some out (keeping ALL of them, you'll need a case of petris) and fruit them individually and you'll see that.
-------------------- Redd Foxx said: If you're offended I don't give a shit and don't come see me no more. Pat The Bunny said: A punk rock song won't ever change the world, but I can tell you about a couple that changed me. bodhisatta said: i recommend common sense and figuring it out. These are the TEKs I use. They're all as cheap and easy as possible, just like your mom.
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