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c10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: Piezo] 1
#23587735 - 08/28/16 05:42 PM (7 years, 5 months ago) |
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if you are worried about "too much work" then definitely avoid isolates lol in the past couple of months i have been through about 800 plates and still dont have any true isolates lol... which is why many of us end up making LI or the like out of good looking clean plates at some point (limited MS) and inoculating with it, fruiting it, and make some clones to play with while we are working on isolates
2. right, thats another fascinating dynamic of MS, all the different aspects which can be influenced by environment, genetics, survivability, etc. I often wonder how much of the same genetics are present in different sectors of a MS plate, and also between fruits of a MS grow. i have started to suspect that some strains are really good at intermingling with other strains, making it very hard to isolate either of them until they split up for some reason, or grow different directions on agar. i know there is a TON of genetic diversity within a MS grow, but i wonder how much genetic similarity there is, ie. how many sectors share strains, and how many fruits are from the same strain and combination of strains. truly fascinating stuff
3. what do you mean by inoculating a jar with a print? one or both of us is confused haha... btw most prints are dirty, rare for them not to be, thats why we use agar to clean up the culture rather than making a dirty spore syringe, etc, and wondering what went wrong when our jars smell sour or turn green. ahhhh rereading it i think i am starting to understand your post a little better, just a little strangely worded. This may be exactly what you are doing, i am just stating it clearly for the benefit of you and anyone working on something similar who might come across the thread
It can actually be tricky sometimes to tell if plates are clean. there is the obvious stuff to look for, but then there is also bacteria that can get down inside the culture in a way that it does not grow separately and does not distinguish itself from the myc on agar. this can be damn tough to catch before jars contam. definitely post pics and we can tell you if we see anything obvious though
but you shouldnt be worried about your genetic choices at this point, thats for sure. really, you havent made any genetic choices yet, since there could be hundreds to thousands of strains present in each initial ms transfer, and you have no way to distinguish one from the other, much less select one. At this point you should just be worried about transferring clean growth to new plates, then let it grow out and do it again, until you are relatively sure its clean. then let a clean plate colonize fully and make a LI, or go agar to grain if you are more patient than me
with MS there really isnt any genetic choice involved, it is 100% luck of the draw
you could of course keep on doing transfers until sectors start to show, and then transfer the aggressive rhizo sectors, and then use that to make LI, and that would likely be more aggressive/rhizo than a less limited MS. but less than 8-10 transfers, it is not really limited at all, just MS.
also bear in mind, you could make an LI from a limited MS rhizo plate and then for some reason the rhizo myc could be totally outcompeted by some tomentose or linear myc that happens to be tangled up in the rhizo myc that was invisible to you during the transfers. there are just so many variables with MS
you could also make a limited MS culture, make a bunch of spawn from it, make 10 monotubs, and have all 10 monotubs turn out ENTIRELY different. some could be albino, some could look like dinosaurs, some could look like pug dogs, but they will most definitely all be different. even the fruits within the same tub will be different, with a potentially high degree of genetic diversity no matter how limited the culture seemed to be, since it was still MS. this is why i say you should not worry about your genetic choices because you really havent made any yet
cloning desirable fruits is definitely the fastest and easiest way to a culture that is more predictable than MS, and more likely to obtain full canopy flushes. take multiple clones and test them separately
the amount of work involved in creating and testing isolates makes it more of a specialized thing, definitely not the easiest. but for those of us who want to collect truly meaningful data points, it is essential
EDIT:
btw i had a question for yall vets:
is there any reason to isolate strains found within a clone? for example, i took several clones from a recent AA+ grow, and they are grown out enough that i could transfer them today. ive noticed several sectors on each clone culture, some rhizo some not so much. is there any reason or benefit in trying to isolate strains from each of these sectors? and if so should they be tested as isolated strains or as subsets of a clone? how do yall usually handle clone plates showing sectors?
im still pretty new to both agar and clones, so dont assume i know anything, assume i am a dumbass and school me
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  C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide "Partial knowledge is more triumphant than complete knowledge; it takes things to be simpler than they are, and so makes its theory more popular and convincing." "Convictions are more dangerous enemies of truth than lies" ― Friedrich Nietzsche
Edited by c10h12n2o (08/28/16 06:06 PM)
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TedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
#23588589 - 08/28/16 10:00 PM (7 years, 5 months ago) |
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In response to Pleze:
2. Unless you are using an isolated strain, yes, you will get various results throughout a grow, especially if it is multi-spore 3. If you are trying to transfer healthy mycelium away from contams then you are using agar for a great purpose 4. Yeah, if you want guaranteed genetics then you will have to isolate a bunch of strains, test them all, and choose 5. Yeah, that's why I was wondering is isolating is generally worth it when you can clone easier. My guess to this problem is clone once you grow and if you want to then start isolating as a side projectI. I just wonder how likely it is to be worth it
-------------------- Alice asked the Cheshire Cat, who was sitting in a tree, “What road do I take?” The cat asked, “Where do you want to go?” “I don’t know,” Alice answered. “Then,” said the cat, “it really doesn’t matter, does it?”
Edited by TedTheHighlighter (08/28/16 10:01 PM)
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MMG
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I'm not too into isolating much so I can't really answer your specific questions on the clone isolation but I would like to tell you that you don't have to fruit out a whole mono to trial different isolates...check out Violets small container grows or pasty's invitro pods too. Doing small grows in containers like the ones shown in these teks makes testing out different isolates for potential winners much easier than doing a bulk grow.
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spacechildo
proletarians rise up


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Re: Strain Isolations on Agar, Pics and Questions [Re: MMG]
#23589072 - 08/29/16 04:32 AM (7 years, 4 months ago) |
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you shouldn't test your cultures on cased grains in cups if you plan to do monotubs with them, they'll behave very differently on cakes,subs,bottles etc.
so you might be throwing away a kickass culture just because you didnt grow it the way it wanted to.
get to know your cultures, what's their likes and dislikes, thats how you can really take advantage.
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c10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: spacechildo]
#23589751 - 08/29/16 11:33 AM (7 years, 4 months ago) |
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thanks for the info MMG!
that was exactly my thought space, very succinctly put
when i first started this project my plan was to test each in its own little bucket a la otto, which has actually been working alright, but i figured since my main method will probably be monotubs for a bit (until i get my automated room built) so i didnt want to miss out on a killer culture that just happens to need a 3" substrate depth or quits the job if we dont give it straw and manure on a trashbag-lined platter
EDIT: here is a pic of one of those buckets btw:

makes sense to test cultures the same way you would ideally fruit them
that said, we are talking a little bit theoretically. mini grows to test isolates could be very useful if someone is limited on space, and i think the odds of finding an isolate that does great in a monotub but sucks on smaller methods is probably VERY small. but since that is one more set of variables i can control for, im gonna be testing in monotubs & bulk trays
Also, for the guy who was talking about cleaning up cultures on agar, here are a few cultures i am in the process of cleaning up, a p. microspora and a cordyceps militaris. these plates are on their second transfer, and are still pretty dirty.

you can see in the cord. m. especially, the line of bacteria all the way down the plate and all the way around in a ring. probably from liquid rolling around. In these, i am taking transfers from the leading edge of the cleanest looking, healthiest myc and transferring to new plates until there is no more visible bacteria, then will make LI. this is the same thing i would suggest doing to clean up a dirty spore print
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  C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide "Partial knowledge is more triumphant than complete knowledge; it takes things to be simpler than they are, and so makes its theory more popular and convincing." "Convictions are more dangerous enemies of truth than lies" ― Friedrich Nietzsche
Edited by c10h12n2o (08/29/16 03:22 PM)
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Piezo
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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
#23589860 - 08/29/16 12:26 PM (7 years, 4 months ago) |
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Quote:
c10h12n2o said: if you are worried about "too much work" then definitely avoid isolates lol in the past couple of months i have been through about 800 plates and still dont have any true isolates lol... which is why many of us end up making LI or the like out of good looking clean plates at some point (limited MS) and inoculating with it, fruiting it, and make some clones to play with while we are working on isolates
Now that I'm pouring agar with stackable PP5 containers, i can do two dozens of plates in one batch. They fit better than plasty plates in my tiny pressure cooker. Short term project is a bulk grow, so isolates will wait.
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c10h12n2o said: 2. right, thats another fascinating dynamic of MS, all the different aspects which can be influenced by environment, genetics, survivability, etc. I often wonder how much of the same genetics are present in different sectors of a MS plate, and also between fruits of a MS grow. i have started to suspect that some strains are really good at intermingling with other strains, making it very hard to isolate either of them until they split up for some reason, or grow different directions on agar. i know there is a TON of genetic diversity within a MS grow, but i wonder how much genetic similarity there is, ie. how many sectors share strains, and how many fruits are from the same strain and combination of strains. truly fascinating stuff
Yes, with this observations, the genome evolution dynamics are also interesting. At the level of a single line of descent, we have haplogroups, groups of genes inherited together. At the "social" level we may have co-evolved strains that grow and sporulate together in carpophores. A bit more than half of the pack needs to sporulate to pass most of the genetic diversity. This only works with the hypothesis of isolated groups, but strains are not species. So if this is real, there must be some mechanisms acting against the genetic drift, like mycelium segregation or the fact that some strains overpower others for a given environment.
I'm also curious about the redundancy of strains in sectored growths. For example, two strains intertwined (A+B) could grow more than two sectors if they separate in a stripes (CW: A, B, A, ..., B). Even with no redundancy, the combined strains could grow in a third sector : (A, A+B, B). With more strains the number of combinations explodes.
I also stand to be corrected here, I'm on my third agar project, and the first two were not really successful...
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c10h12n2o said: 3. what do you mean by inoculating a jar with a print? one or both of us is confused haha... btw most prints are dirty, rare for them not to be, thats why we use agar to clean up the culture rather than making a dirty spore syringe, etc, and wondering what went wrong when our jars smell sour or turn green. ahhhh rereading it i think i am starting to understand your post a little better, just a little strangely worded. This may be exactly what you are doing, i am just stating it clearly for the benefit of you and anyone working on something similar who might come across the thread
Sorry about that, not native, still learning.  Yes, I'm currently using agar to clean MS culture, but considering to try MS in small cakes for making clones directly. As said by MMG, it should be possible to contain contaminations with isolated growth (bottle teks). Then the clone is passed through some plates to clean it or to isolate strains.
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c10h12n2o said: with MS there really isnt any genetic choice involved, it is 100% luck of the draw
you could of course keep on doing transfers until sectors start to show, and then transfer the aggressive rhizo sectors, and then use that to make LI, and that would likely be more aggressive/rhizo than a less limited MS. but less than 8-10 transfers, it is not really limited at all, just MS.
Maybe MS mycelium do less sectoring than clones because its strains weren't subjected to bulk grow and fruiting ? I have not seen rhizomorphic growth yet in my limited MS plates. It's seems easier to get rhizomorphic from clones. Plasticity and/or selection ?
As you said, changes in characters also happen with liquid inoculants. The environmental pressures from the spore germinations until fruiting in bulk could be decisive on the results, maybe more than the original set of genetics. That's the exciting part in getting contaminations and see them eaten by the cubensis mycelium.
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c10h12n2o said: Also, for the guy who was talking about cleaning up cultures on agar, here are a few cultures i am in the process of cleaning up, a p. microspora and a cordyceps militaris. these plates are on their second transfer, and are still pretty dirty.

you can see in the cord. m. especially, the line of bacteria all the way down the plate and all the way around in a ring. probably from liquid rolling around. In these, i am taking transfers from the leading edge of the cleanest looking, healthiest myc and transferring to new plates until there is no more visible bacteria, then will make LI. this is the same thing i would suggest doing to clean up a dirty spore print
Ok thanks for the illustrated example.
Practical question : sometimes i transfer aerial mycelium with a needle without touching the agar surface. It's a bit tricky to peel of the mycelium from the needle, but I think it transfer less bacteria. Am i the only one doing this ?
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c10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: Piezo]
#23595825 - 08/30/16 10:40 PM (7 years, 4 months ago) |
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Quote:
Piezo said:
Now that I'm pouring agar with stackable PP5 containers, i can do two dozens of plates in one batch. They fit better than plasty plates in my tiny pressure cooker. Short term project is a bulk grow, so isolates will wait.
if i am not mistaken, the definition of a pasty plate" would be using commonly available PP5 containers and no-pour agar instead of petris (which arent available to some people) and pouring (which some people either struggle with or would rather avoid), i think thats the main idea he was trying to introduce people to with that tek. (your stackable pp5 containers are probably still pasty plates)
But for what you are trying to do, i would suggest doing what i did: streak some plates with spores, transfer healthy myc to new plates, let it grow out a bit, and either do a few more transfers or go ahead and make a LI (liquid inoculant) using the fresh plate once it is colonized a bit. then use the LI however you like, and clone some fruits. after that, isolation work is a matter of preference, sounds like clones would be best suited for your purpose
Quote:
Maybe MS mycelium do less sectoring than clones because its strains weren't subjected to bulk grow and fruiting ? I have not seen rhizomorphic growth yet in my limited MS plates. It's seems easier to get rhizomorphic from clones. Plasticity and/or selection ?
something like that lol... the first few plates in a series of isolation transfers (starting with MS) are almost never rhizomorphic in my experience. it is mainly a matter of the jungle of strains present growing on, through, between, and beside each other. I am not sure why such large numbers of strains group together at this point and form circular growth (rather than, say, 400,000 little sectors), where a smaller number of strains would likely show clear sectoring
I am working on isolating strains from several races. i started out with 4 races of cubes and 1 pan, and have since started 4 more races. out of these, only 2 races have shown rhizomorphic growth at all, even 12 transfers in most of them are either linear growth or tomentose, and often perfectly round and maybe a few cells thick, just wimpy.
on the other hand, by about the 4th or 5th transfers, those some of the plates from those 2 races started showing rhizo sectors, and i started focusing on transferring those. after a few more transfers the plates in those series were very aggressive with several rhizo sectors each, totally different than the type of growth i was seeing on the earlier plates and on other races i am working on
when i started using these very rhizomorphic plates to make LIs, my colonization times went from weeks to days (one of these monotubs literally colonized in 4 days, where my initial monotub of the same race took about 2 weeks). if i had only been working with a few plates/races, i might have never gotten rhizo growth, or i might just not have gotten it yet. you might have to transfer a hundred times just to see clear sectors :/ thats why it makes sense to dilute spores
the reason why it is easier to get rhizo growth from clones is probably because of the MUCH smaller number of total strains present in the culture/biopsy
i think you are severely underestimating the number of strains present (as i did, and might still be)
Quote:
That's the exciting part in getting contaminations and see them eaten by the cubensis mycelium.
i agree, they are fun to watch, i used to keep every contammed sample just to watch it.
BUT make sure you separate it from your serious projects, and dont get attached to the jars/bags/dishes.
Focus on learning how to obtain clean cultures and figuring out the point of failure and resolving it, that would benefit you the most in the long run. you want to be sure a culture/spawn is 100% clean before using it for g2g, or betting on it for anything serious. personally, i want to do everything i can to avoid having a tub full of rotten feet smell, and clean spawn is the least we can do to ensure success
Quote:
Practical question : sometimes i transfer aerial mycelium with a needle without touching the agar surface. It's a bit tricky to peel of the mycelium from the needle, but I think it transfer less bacteria. Am i the only one doing this ?
why would you do that? i did it a few times, in my early plates where i wanted to see rhizo myc soooooo bad but there just wasnt. any. there.
i was able to fool myself into thinking "hey, there in the middle, that looks rhizomorphic! that big fuggin thing coming out of the top of the agar i transferred!"... but i think i was bullshitting myself. there was no rhizo growth on those plates
what there was was a piece of retarded myc that grows straight in the only direction without agar. aerial myc is not a good quality, certainly not a feature you should try to transfer.
also, if you are doing what i was doing, then you are probably grabbing aerial myc that is very near the origin of the dish's growth, maybe even growing off of the agar you transferred, not the agar you transferred to. if this is the case (as it was for me), you are misunderstanding what you are trying to accomplish by using agar. the whole point is to have controlled growth across the surface of the agar so that you can distinguish various characteristics and transfer desirable sectors. if you are pinning growth from your initial transfer, that did not even grow across the agar at all, it throws away all the advantages of using agar for this purpose
at least that is what made sense to me, so i stopped doing that
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  C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide "Partial knowledge is more triumphant than complete knowledge; it takes things to be simpler than they are, and so makes its theory more popular and convincing." "Convictions are more dangerous enemies of truth than lies" ― Friedrich Nietzsche
Edited by c10h12n2o (08/30/16 10:42 PM)
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MMG
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Re: Strain Isolations on Agar, Pics and Questions [Re: spacechildo]
#23595880 - 08/30/16 11:03 PM (7 years, 4 months ago) |
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I see your point there. bottle growing aren't just cased grains, when I do bottles I spawn at around 1:5 or so ratio (spawn to sub) mixing the spawn together with the bulk sub (cvg) and then I case them. You can eliminate some of the unknowns or type 1 errors (discarding a culture given that it was a good one) by doing multiple bottles, say 4 or 5 per isolate. Still less hazzle than doing a mono and it will give you a good idea of how the culture responds. if 3 or 4 out of the 5 show good results then you can say with a good probability that it's a keeper or one to keep testing on. TBH I hadn't heard of mushrooms being picky over how much sub they're fruiting on. types of sub, and growth parameters yes (but space - no) if you're doing the same cvg sub for bottles or a mono (keeping the sub the same) then I don't see how it would affect performance given that fruiting parameters are kept on check, illuminate me if I'm wrong here. edit: sub depth probably does play some role in the development (for example pans liking more shallow subs) but in general for cubes I don't think it would matter much (a good culture will perform well regardless), again this is my 2c
edit 2: at OP - that's so cool that you're cultivating p. microspora, do you plan on using it to digest plastics or just doing it for the novelty?
Edited by MMG (08/30/16 11:41 PM)
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c10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: MMG]
#23598450 - 08/31/16 06:17 PM (7 years, 4 months ago) |
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great info MMG 
i actually just harvested a otto bucket today, one HUGE cluster! 

im about to clone one of those monsters that poked its head out
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  C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide "Partial knowledge is more triumphant than complete knowledge; it takes things to be simpler than they are, and so makes its theory more popular and convincing." "Convictions are more dangerous enemies of truth than lies" ― Friedrich Nietzsche
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MMG
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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
#23598485 - 08/31/16 06:28 PM (7 years, 4 months ago) |
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damn! that's a nice cluster right there. is this one of the isolates you've been working on or MS? looks like the hard work is paying off!
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c10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: MMG]
#23598554 - 08/31/16 06:44 PM (7 years, 4 months ago) |
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sorry, i forgot to answer your question about p. microspora: yes, i have various projects in mind! so excited about that one. i know it is stupid, but i have been fantasizing about making it saltwater tolerant, or building some kind of platform that would allow it to digest some of those plastic islands in the ocean... i know its dumb for at least a dozen reasons, but its a thought haha... also plan to experiment with various trash-to-food projects 
thanks buddy!! that bucket is actually one of the first things i did when i started this project, was planning to work out a good system for testing isolates and figured i would practice with MS and clones. the plan was to use these little pails with 1qt of spawn, and fruit them like otto or monos. they ended up taking a lot longer to colonize than i had hoped, but this AA one finished faster than the others
so no, that bucket is just MS, LI wasnt even really made from a good looking plate if i recall... i cant wait to start fruiting some of the AA tubs i spawned 5 and 6 days ago, they are looking GREAT! much more rhizo, much more limited genetics, and the monotubs were solid white in 5 days!!! crazy stuff.. i almost fruited them today, but there is one tiny spot on each that is still a little uncolonized, about the size of a pinhead, and i figure letting it colonize another day or two certainly wont hurt
I plan to make 4 big spawn bags today from an AA clone culture from my last monotub, and i am about to clone one of those monsters in that giant cluster
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  C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide "Partial knowledge is more triumphant than complete knowledge; it takes things to be simpler than they are, and so makes its theory more popular and convincing." "Convictions are more dangerous enemies of truth than lies" ― Friedrich Nietzsche
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TedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
#23599114 - 08/31/16 09:18 PM (7 years, 4 months ago) |
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Quote:
c10h12n2o said: sorry, i forgot to answer your question about p. microspora: yes, i have various projects in mind! so excited about that one. i know it is stupid, but i have been fantasizing about making it saltwater tolerant, or building some kind of platform that would allow it to digest some of those plastic islands in the ocean... i know its dumb for at least a dozen reasons, but its a thought haha... also plan to experiment with various trash-to-food projects 
Doesn't sound dumb to me! I think it would be awesome for you to dig into what you can do in mycology, especially if you look into beneficial stuff like your ideas
-------------------- Alice asked the Cheshire Cat, who was sitting in a tree, “What road do I take?” The cat asked, “Where do you want to go?” “I don’t know,” Alice answered. “Then,” said the cat, “it really doesn’t matter, does it?”
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MMG
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I second that, no idea that is pro-world/life is dumb in my book! speaking about remediation I wanna restart my efforts on decomposing cigarette butts (cellulose acetate fiber) with oysters. I plan on "training" the myc by slowly exposing it to the filter material (along with grain) until I secure a good candidate for the application...one of the things I was debating was the feasibility of using the mushroom mycelium directly on dirty filters which is probably null (proper sterilization is most likely necessary) so I don't know how cost-efficient or sustainable it would be. It'd have to be like kilos of cigarette butts to justify the sterilization but it might be worthwhile...guess I'll have to get to work on it and report back! so wait, the decomposed material from the P. Microspora is edible? that's pretty crazy haha in the case of the cig butts, given that they're carcinogenic (a tleast when utilized) the mushrooms fruits (if any are produced at all) are non edible or deemed unsafe for human consumption. I'd be happy to just decompose the waste at a much faster rate than it does in landfill or even worse in the ocean for the case of cig waste. then composting it of course back on topic...@op, that's the thing with isolates man, sometimes (at least in my case) isolating yielded worse or basically the same results that an MS culture did so I haven't done too much isolating because for my purposes I don't see it as a necessity. if it's clean I'm happy and if I get a big ass cluster like the one you did then awesome, just clone some tissue from it and isolate a bit from there and re-test again to see how it performs.
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TedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: MMG]
#23600110 - 09/01/16 06:45 AM (7 years, 4 months ago) |
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@MMG Sounds like a great idea. Report back on that project if you start it up
-------------------- Alice asked the Cheshire Cat, who was sitting in a tree, “What road do I take?” The cat asked, “Where do you want to go?” “I don’t know,” Alice answered. “Then,” said the cat, “it really doesn’t matter, does it?”
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TedTheHighlighter
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So does weetie have any updates for us?
-------------------- Alice asked the Cheshire Cat, who was sitting in a tree, “What road do I take?” The cat asked, “Where do you want to go?” “I don’t know,” Alice answered. “Then,” said the cat, “it really doesn’t matter, does it?”
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Supalemonhaze
Spore syringe hater.



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Quote:
TedTheHighlighter said: So does weetie have any updates for us?
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TedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: Supalemonhaze]
#23637272 - 09/11/16 09:53 PM (7 years, 4 months ago) |
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Does the OP have updates?
-------------------- Alice asked the Cheshire Cat, who was sitting in a tree, “What road do I take?” The cat asked, “Where do you want to go?” “I don’t know,” Alice answered. “Then,” said the cat, “it really doesn’t matter, does it?”
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c10h12n2o
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hey guys sorry for the late reply, had a small series of minor emergencies come up with work and family and health, been trying to get caught up. meant to update this a while back, i will post a bunch of relevant pics shortly as soon as i have time to organize em
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MMG said: so wait, the decomposed material from the P. Microspora is edible? that's pretty crazy haha in the case of the cig butts, given that they're carcinogenic (a tleast when utilized) the mushrooms fruits (if any are produced at all) are non edible or deemed unsafe for human consumption.
yep, apparently they are! here is a page where someone has made a really cool little self contained system. it actually looks pretty sweet, a very different approach than most projects ive seen

also, i am curious about the oysters picking up toxins, i know they are really good about concentrating heavy metals (or bad, depending on whether you are trying to remove heavy metals from a substrate or eat mushrooms), but in Mycellium Running Stamets mentions a project where the government was testing several ways to clean up oil spill type stuff, testing some bacteria, algae, and a few other means of breaking down the hydrocarbons. Stamets said that at the end of the trial, the other piles were rotten, but the oyster pile had produced a mountain of beautiful fruits that were safe to eat. between concentrating heavy metals, and what you are saying about cigarette toxins, it seems amazing (or questionable) that Stamets petro oysters were really safe to eat
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MMG said:back on topic...@op, that's the thing with isolates man, sometimes (at least in my case) isolating yielded worse or basically the same results that an MS culture did so I haven't done too much isolating because for my purposes I don't see it as a necessity. if it's clean I'm happy and if I get a big ass cluster like the one you did then awesome, just clone some tissue from it and isolate a bit from there and re-test again to see how it performs.
oh goodness! im worried about exactly that! i often wonder if a well organized culture of compatible strains might perform than certain isolates. i guess it just underscores the need for extensive testing and good notes huh
I did take several clones that are lookin good I have about 10 i am working with right now, isolating some out further, and have 2 particular clone cultures currently colonizing tubs, and a TON of grainspawn colonizing from 2 different clone cultures. one of the clone cultures ive inoculated with comes from one of those huge clusters, the other from a pin, and about 8 others im isolating further. Im really excited about testing some of the clones and isolates!! i look forward to testing clone cultures that have most of the strains present in the initial biopsy vs those that have been further isolated (the rhizo sectors)
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TedTheHighlighter said: Does the OP have updates?
sorry for the delay my friend... life got a little crazy! but i have stayed on top of my projects though
I have been through a little over 1000 plates since i began this project, and boy have i learned a lot + destroyed a few of my delusions hahaha...
when weetsie started his project, i did the same thing, 20 different initial plates, with varying amounts of spores on each one (some had a ton like weetsies, some had far less, some were streaked, some were dilluted), using 3 races of cubes and one p. sub..
Out of all 20 different series, i dont think any of them are anywhere near an isolate, even as we speak. The different series are on between their 9th and 13th transfers, and there are multiples of quite a few from each generation after 3rd. RR might be able to get isolates in just a couple of transfers, but apparently i have a LOT of learning left to do before i can pull off something like that haha....
it has been an informative experience though, thats for sure. like someone mentioned at various points before (i think it was bodhi and supalemonhaze and space), sometimes you will have a VERY rhizomorphic culture that looks like there could be "at most 1-3 strains present", but you would be totally wrong. made me want to scream the first dozen or so times i transfered a tiny little sliver from what i believed would contain only 1 highly rhizo strain, only to have it turn completely tomentose and have a 8+ sectors on the next plate
I have also been working with lots of clone cultures in addition to the isolations im working on, and they have been fascinating. I expected to see 1-3 sectors in clones, but to my surprise, there are often many more. and when i transfer a rhizo sector from a clone culture, expecting it to be an isolate, it often turns cottony, or shows many previously unseen sectors
so yeah, every time i have thought i had an isolate, further transfers have revealed that there were many, many strains present
I will update with a bunch of pictures, from different parts of the process, and will post updates as i get closer to isolates
lol hopefully weetsie is having better luck than me, and if so i hope he enlightens me!! i appreciate everyone who has contributed info and advice, you guys rock
btw, i think it was supalemonhaze who told me that trying to work with isolating multiple races at the same time is a hellacious amount of work, and BOY WAS HE RIGHT. this project quickly became like a job, and now that i am working with 7 races of cubes and 6 other species, it is downright ridiculous. ive been keeping up to the best of my ability, but i certainly would not recommend working with this many different races at a time, even if you do have a ridiculous work ethic, it ends up being an insane amount of work (and if you just want predictable crops the same end can be achieved a lot easier; me personally though, i cant wait to start collecting some meaningful datapoints testing isolates)
EDIT: BTW, i meant to ask, do yall have any good tips for disposing of old plates? i am cleaning up, and have 2 black trashbags full of plates that are from earlier in these transfers. that is a TON of plates, most are clean but i just have no use for them, since i am much later in these isolation processes now. it seems like a VERY sketchy 2 bags of trash, that many labeled petri dishes might get the wrong kind of attention if it was ever noticed.
obviously, would want to make sure no mail or anything with my name, address, etc is in there with it, and dispose of it at a random dumpster, id burn it if i wasnt in a city. i usually throw away small amounts of sketchy stuff through regular trash pickup, but this many dishes looks seriously sketchy. any advice?
much obliged my friends
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  C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide "Partial knowledge is more triumphant than complete knowledge; it takes things to be simpler than they are, and so makes its theory more popular and convincing." "Convictions are more dangerous enemies of truth than lies" ― Friedrich Nietzsche
Edited by c10h12n2o (09/19/16 01:46 AM)
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TedTheHighlighter
Cheshire Cat


Registered: 12/09/14
Posts: 490
Last seen: 2 months, 7 days
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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
#23658046 - 09/19/16 07:52 AM (7 years, 4 months ago) |
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Glad to see you haven't given up, in fact you've done just the opposite. You are definitely proving that it takes many transfers to reach isolation. I really do wonder how RR did it with just a couple transfers. I would definitely love to hear about your cloning results along with isolation results. I am still very curious about clones vs isolates
-------------------- Alice asked the Cheshire Cat, who was sitting in a tree, “What road do I take?” The cat asked, “Where do you want to go?” “I don’t know,” Alice answered. “Then,” said the cat, “it really doesn’t matter, does it?”
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Wasn't this the thread we were supposed to be proven wrong that isolating takes like 5 transfers or some BS? What's up with that shit?
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