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3SIXTY5
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Clone --> Isolate, senescence?
#26908137 - 08/30/20 03:14 PM (3 years, 5 months ago) |
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Hey, so I grew some PE6 out on BRF, put a clone to agar, and started transferring, U use 100mm dishes and I always transfer once the culture has colonized about 40%-60% of the dish. I'm currently on my 8th transfer and my last bulk run with my T7 dishes all contaminated. That got me thinking, is t possible for a cloned culture to reach senescence before you even reach an isolated set of genetics?
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Forrester
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908348 - 08/30/20 04:43 PM (3 years, 5 months ago) |
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I don't think so. But if it's looking tired, switch up your agar recipe to get some different nutrients into it. Sometimes running across a couple different recipes will even get an old, tired culture going again.
-------------------- Repugnant is a creature who would squander the ability to lift an eye to heaven, conscious of his fleeting time here. ------------------- Have some medicinal mushrooms and want to get the most out of them? Try this double extraction method.
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jcm4620
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Re: Clone --> Isolate, senescence? [Re: Forrester]
#26908367 - 08/30/20 04:54 PM (3 years, 5 months ago) |
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u should be making masters of each transfer and fruiting them all out so you can go back and slant or transfer out further if wanted to narrow it down even more. while keeping good performers and junking the rest. cuz the way u make it sound is that u just keep pulling transfers off the tissue sample plate each time and what ever your fruiting out your just basicly losing them as you go.
thats what it sounds like ur saying to me at least so if im wrong im sorry.
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: jcm4620]
#26908443 - 08/30/20 05:33 PM (3 years, 5 months ago) |
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Quote:
jcm4620 said: the way u make it sound is that u just keep pulling transfers off the tissue sample plate each time
That's exactly what I'm doing, I'm only interested in single sets of genetics. I cloned that sample just to narrow down the genetics before I started isolating, because when I start from spore I find reaching an isolate just takes too long, I'm hoping that I can just take a shortcut to an isolate. as I understand it, starting from spore will give me the best shot at an amazing isolate, but that's going to take too long, so I'm doing it this way to look for something that's pretty damn good, maybe not absolutely amazing, but I don't need that. I mainly want one that's going to pin and grow to maturity at basically the same rate. I'll make masters of the isolates, but that's it. I'm growing them out as I go just to get some along the way.
Edited by 3SIXTY5 (08/30/20 05:34 PM)
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: Forrester]
#26908465 - 08/30/20 05:42 PM (3 years, 5 months ago) |
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Quote:
Forrester said: I don't think so. But if it's looking tired, switch up your agar recipe to get some different nutrients into it.
Yeah, it's still growing out fine on dishes I think, they're not in an incubator so it's a little harder to tell with the weather fluctuation so much lately. So could it still grow alright on dishes, and grains, but once it hits substrate it's too tired?
I probably fucked up my bulk subs. I was just thinking maybe it was the culture getting weaker and that's why my subs contaminated, I just didn't want to do another grain spawn and substrate run before I find out if it's the culture and not my substrate process that fucked me Up.
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Roger Clemency
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908472 - 08/30/20 05:45 PM (3 years, 5 months ago) |
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What was your bulk sub recipe and prep?
-------------------- Sour grapes, sweet revenge Heaven starts right where hell ends
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trubblesome
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908477 - 08/30/20 05:48 PM (3 years, 5 months ago) |
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Quote:
3SIXTY5 said:
Quote:
jcm4620 said: the way u make it sound is that u just keep pulling transfers off the tissue sample plate each time
That's exactly what I'm doing, I'm only interested in single sets of genetics. I cloned that sample just to narrow down the genetics before I started isolating, because when I start from spore I find reaching an isolate just takes too long, I'm hoping that I can just take a shortcut to an isolate. as I understand it, starting from spore will give me the best shot at an amazing isolate, but that's going to take too long, so I'm doing it this way to look for something that's pretty damn good, maybe not absolutely amazing, but I don't need that. I mainly want one that's going to pin and grow to maturity at basically the same rate. I'll make masters of the isolates, but that's it. I'm growing them out as I go just to get some along the way.
you already have what you're looking for. transfers from clone plates are to clean up from contams. The genetics are just that - clones, they're narrowed down to just one set.
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jcm4620
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908479 - 08/30/20 05:49 PM (3 years, 5 months ago) |
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there is no such thing as a shortcut to a true isolate tho. thats not technically an isolate there is a fuck ton of sets of genetics in every transfer you make. all your doing is just taking transfers from a clone. the term isolate is actually used rather loosely by people. just cuz you think your "isolating" one piece of myc dosent make it a true isolate. its far to much information to really explain typing with my thumbs on my phone but if your interested in learning about true isolates and mono and dikaryotic cultures there is plenty of info available. all i was getting at with what you are doing is potentially throwing away your best genetics from your clone since your not taking transfers and making a master to go back to if it ends up being a great fruiter is all i was saying. but no matter how many transfers you make you really still just have a clone
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jcm4620
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Re: Clone --> Isolate, senescence? [Re: jcm4620]
#26908482 - 08/30/20 05:51 PM (3 years, 5 months ago) |
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i think jake actually has shit on hear explaining actual isolates very well that dudes the fucking man when it comes to that shit so if u search it toss his name in there also
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: jcm4620]
#26908517 - 08/30/20 06:10 PM (3 years, 5 months ago) |
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Quote:
jcm4620 said: i think jake actually has shit on hear explaining actual isolates very well that dudes the fucking man when it comes to that shit so if u search it toss his name in there also
I'll definitely check that out, thanks.
I understand it's not a true isolate, maybe I'm using the wrong terminology, maybe I don't understand the process, but how can a clone be one set of genetics if its still sectoring on a plate? Also I remember reading a post by RR explaining that clones aren't usually single sets of genetics.
I understand monokaryotic mycelium is formed when one spore is germinated and that it's a non fruiting strain without another monokaryon. Is this related to an isolate?
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: trubblesome]
#26908522 - 08/30/20 06:13 PM (3 years, 5 months ago) |
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Quote:
trubblesome said: you already have what you're looking for. transfers from clone plates are to clean up from contams. The genetics are just that - clones, they're narrowed down to just one set.
Really, I've read otherwise, I remember an RR post one clone being made up of multiple sets of genetics.
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jcm4620
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908538 - 08/30/20 06:20 PM (3 years, 5 months ago) |
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clones have more genetics than i would ever attempt or want to try to count lol
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jcm4620
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Re: Clone --> Isolate, senescence? [Re: jcm4620]
#26908558 - 08/30/20 06:28 PM (3 years, 5 months ago) |
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when working with clones your really looking for traits more than the actual genetics themselves. but yet when u have a good clone its common and often refered to as good genetics lol i know right 😂words are fun arent they😃
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: jcm4620]
#26908596 - 08/30/20 06:51 PM (3 years, 5 months ago) |
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Quote:
jcm4620 said: clones have more genetics than i would ever attempt or want to try to count lol
Yes I hear you on that, but my understanding was that when one drop of spore solution or one streak of a loop would have more sets of genetics than a cloned tissue sample from a fruit body, am I wrong about this?
I'm not looking for a true isolate or I would have started from spore, I'm looking for a single set of genetics so the whole substrate will pin and reach maturity within 24 hrs or so, so I don't have a substrate taking up space in my fruiting chamber for too long. Once I have a bunch of single genetic dishes, then I will test them for yield and all that.
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trubblesome
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Re: Clone --> Isolate, senescence? [Re: jcm4620]
#26908601 - 08/30/20 06:53 PM (3 years, 5 months ago) |
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Do you have a source on that? I've never read that anywhere and it doesn't make a lot of sense intuitively based on everything I have read about the life cycle of mushrooms. As I understand, in the flesh of the fruiting body the cells should all be identical and dikaryotic, containing the unfused haploid nuclei from the hyphae that paired and formed the primordia. in the basidia the nuclei fuse to create a diploid set and then undergo meiosis to create haploid spores. I looked for something myself and all I could find was an article about some Armillaria species undergoing a haploidization -> diploidization -> haploidization before producing primordia but that seemed to be unique to them and perhaps a reason for their ability to grow large and old.
my intuition says that differences in fruiting body phenotype would be due to conditions and available resources, not genetics.
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3SIXTY5
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Quote:
Roger Clemency said: What was your bulk sub recipe and prep?
SUBSTRATE PREP
I sterilize my bulk substrate in a sort of modified insulated bucket Tek style bin, I boil water in the bin, wait until it's over 200f, dump in my bulk ingredients (780 grams coco + 225 grams verm + a handful of gypsum + a little more than 3 litres of water, per 5lb bag spawn) let it sit for about 2 hours, mix, let it sit for another 2 hours or so, then mix again. The bin stays above 180f for like 12-18hours, I usually have to keep mixing and waiting for 2-3 days before the substrate is back under 100F.
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: trubblesome]
#26908616 - 08/30/20 07:03 PM (3 years, 5 months ago) |
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Quote:
trubblesome said: Do you have a source on that?
RogerRabbit said: Yes. More than one strain can be present in an individual fruit. Thus, a clone is not an isolated strain, unless it was grown from an isolated line of mycelium. RR
I can link you the post if you want, pretty interesting, but a long read.
Edited by 3SIXTY5 (08/30/20 07:11 PM)
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trubblesome
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908645 - 08/30/20 07:27 PM (3 years, 5 months ago) |
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yeah, that would be great, thanks
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3SIXTY5
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Re: Clone --> Isolate, senescence? [Re: trubblesome]
#26908646 - 08/30/20 07:28 PM (3 years, 5 months ago) |
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trubblesome
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Re: Clone --> Isolate, senescence? [Re: 3SIXTY5]
#26908689 - 08/30/20 07:57 PM (3 years, 5 months ago) |
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if he's just pointing to sectoring on the plate as evidence that the mycelium is not actually genetically identical, I don't know, i'm not super convinced.
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