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InvisibleMunchauzen
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Re: Strain Isolations on Agar, Pics and Questions [Re: Rooster Cogburn]
    #23576120 - 08/25/16 01:34 PM (7 years, 6 months ago)

That's a pretty big 'if' thou. Good on him for trying but I give it a pretty slim chance. prove me wrong, plz.

Edited by Munchauzen (08/25/16 07:43 PM)

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Invisibleweetsie
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Re: Strain Isolations on Agar, Pics and Questions [Re: Munchauzen]
    #23576354 - 08/25/16 02:56 PM (7 years, 6 months ago)

All the plates used so far:





One of the transfers resulted in a very circular cottony growth which I don't plan to continue with. The other has a large rhizomorphic sector which I made two more transfers from.

A photo of said plate with the lid off:






Quote:

Munchauzen said:

still, dont know why he is waiting for it to grow out to the edge of the plate to make a transfer. Weetsie, stop pussy-footin' around and get crackin'!




Because the goal is to do it in as few transfers as possible.

The rhizomorphic mycelium I'm after takes time to develop, if I transferred too soon I might not see a rhizomorphic sector or any sectoring after the first transfer.

The wedge I transfer will also be relatively smaller the more I let the plate grow out which I think is key to what I'm trying to do.

I imagine letting the plate grow out more will also thin the heard so to speak.


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OfflineTedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: weetsie] * 1
    #23576475 - 08/25/16 03:31 PM (7 years, 6 months ago)

This thread is bangin


--------------------
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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OfflineKenetic
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Re: Strain Isolations on Agar, Pics and Questions [Re: TedTheHighlighter]
    #23576488 - 08/25/16 03:34 PM (7 years, 6 months ago)

I think even if you took a small chunk you'd still get tons of genetic diversity.


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InvisibleMunchauzen
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Re: Strain Isolations on Agar, Pics and Questions [Re: Kenetic]
    #23577463 - 08/25/16 07:44 PM (7 years, 6 months ago)

I guess we'll see

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Invisiblec10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: Munchauzen]
    #23578444 - 08/26/16 02:37 AM (7 years, 6 months ago)

Quote:

spacechildo said:
Quote:

c10h12n2o said:
1) its perfectly symmetrical to the agar i transferred though, and using calipers to measure from the edge of the transferred agar to the leading edge of the myc it is almost dead even all the way around. this plate is either 11 or 12 transfers in

2) btw i had been meaning to ask you something spacechildo: when you have a plate that is heavily sectored showing 2 distinct kinds of myc, and there are 3-4 sections of aggressive rhizo myc and 3-4 linear sectors, and some or all of the rhizo sectors look similar, or are even connected, do you transfer all the good looking sectors, even though there is a good chance that some of them are the same? especially late into a series of transfers, like 10-15th plate.

3) sometimes lately it has seemed like i am transferring the same thing twice on the same plate, what appears to be 4 sectors might only be 1 "sector" that is on top/under another sector that is diving it (not to say that this single "interrupted sector" is an individual strain, of course there could be numerous compatible strains forming the same "sector")

4) i suppose there is no harm in labeling and testing them all separately, but im working on so much right now that i would like to minimize wasted time/space on testing strains X, Y, and Z against each other only to discover/suspect that they are indeed only one strain.

let me know if i need to clarify anything. i would really like to hear your thoughts on this and potential ways to negate this possibility without missing out on potentially valuable strains

warm regards




1, well try just 1 more transfer and make it real small, I still doubt it but I've been wrong before :shrug:

2, just look for fast growing strong looking myc. all your plates seem nice and healthy so just pick the fastest growing sectors as that's the only trait you can see on agar.

3, that's confusing :lol: its perfectly normal that sectors grow together and what looks like 1 is really 10 which you'll find out later on as you keep transferring.

4, yeah that's a bitch, in a perfect world you'd test them all but :shrug: do the amount of searching you can/have time for.




Makes sense, thanks man. I will post a pic tomorrow of some plates that show the kind of growth that makes me wonder about that, just for talking points

Quote:

Munchauzen said:
still, dont know why he is waiting for it to grow out to the edge of the plate to make a transfer. Weetsie, stop pussy-footin' around and get crackin'!




Yo munch, can you elaborate on why there is no need to let it grow out? I'm curious if I'm misunderstanding something, but shouldn't letting it grow out allow more chance for sectors to show, and give more opportunity for strains to outpace each other? I usually end up making transfers about every 3-5 days or as soon as I see something sexy, but if the goal is to get an isolate in the fewest possible transfers, why wouldn't he want to give strains time to outpace each other?

I for one appreciate weetsie's experiment and think it is right on topic for the thread, will certainly be useful to anyone with similar questions who reads in the future

Wow blindingleaf, killer plate, beautiful. Did you start with diluted spores?

Also, I have some pan tropicalis on agar right now about 8 transfers in, looks far whisper though, I will post a pic tomorrow

Edit: looking nice weetsie!! Fun experiment, very helpful for readers regardless of "proving" anything

Edited by c10h12n2o (08/26/16 02:39 AM)

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Invisibleblindingleaf
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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
    #23578582 - 08/26/16 04:51 AM (7 years, 6 months ago)

no, just normal spore swipe.

it was just random that it happened in 8 transfers..its not something I expect.

I let mine grow out too once I'm on later plates, like 3-5+ xfers.  not all the way to the edge, but about the size of that picture.  I don't know if it technically helps, but i personally find it easier to spot where I wanna pull from later rather than sooner.


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OfflineTedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: blindingleaf]
    #23579152 - 08/26/16 09:45 AM (7 years, 6 months ago)

Whats the advantage of using a spore swipe rather than a syringe?


--------------------
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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Offlinespacechildo
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Re: Strain Isolations on Agar, Pics and Questions [Re: TedTheHighlighter]
    #23579164 - 08/26/16 09:50 AM (7 years, 6 months ago)

not having excess water on your plate. squirt some spore water on the inoc-loop if you only have syringes. or just "spill" some on the tip of the needle and swipe that.

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OfflineTedTheHighlighter
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Re: Strain Isolations on Agar, Pics and Questions [Re: spacechildo]
    #23580207 - 08/26/16 02:01 PM (7 years, 6 months ago)

Okay, here's another noob question
Whats the advantage in swiping the spores in a "Z" or "L" shape?


--------------------
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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InvisibleInocuole
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Re: Strain Isolations on Agar, Pics and Questions [Re: TedTheHighlighter] * 1
    #23580238 - 08/26/16 02:10 PM (7 years, 6 months ago)

Z is for potency and L is for colonization speed.


Not really, it's just to spread them evenly.  Increases the chances of clean germination by giving a wider area for the (hopefully few) contaminants to be separated from the myc.


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Re: Strain Isolations on Agar, Pics and Questions [Re: Inocuole]
    #23580474 - 08/26/16 03:24 PM (7 years, 6 months ago)

Ahhhhh thanks for the responses


--------------------
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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Invisiblec10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: TedTheHighlighter]
    #23580572 - 08/26/16 03:45 PM (7 years, 6 months ago)

here is a killer video explaining how to streak plates for isolating bacterial colonies:


obviously we are working with fungus instead, but same principles apply.

i have started doing it this way, it is VERY helpful for isolating away from contaminants in a syringe or print, and also useful for having literally thousands of potential colonies to choose from on your initial plate


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C10's Agar Guide + Tips and Tricks | c10's Flow Hood Build Guide


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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
    #23582212 - 08/27/16 12:12 AM (7 years, 6 months ago)

something that hasn't been mentioned in this thread but I have heard others mentioned when isolating agar is the size of the agar plate.
those 110mm plates are specially good (IMO) for isolating because given that you give the mycelium enough space to colonize the easier it will be to pick out different sectors so IMO it should be possible to achieve an isolate in less than 10 plates or so (specially the big ones)... just my 2c
*flame shield up* :heart:


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OfflineKenetic
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Re: Strain Isolations on Agar, Pics and Questions [Re: MMG]
    #23582515 - 08/27/16 04:57 AM (7 years, 6 months ago)

I just got a bunch of 60mm ones.  Should be easier to pour in a GB.


--------------------
Todo Cambia
   

               
                                                :cookiemonster::elmo:



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Invisiblec10h12n2o
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Re: Strain Isolations on Agar, Pics and Questions [Re: Kenetic]
    #23583335 - 08/27/16 12:06 PM (7 years, 6 months ago)

Good point MMG, makes sense to me, i need to get some variety rather than just the standard 90mm

Even more to the point i was asking munch, why he said there is no need to let them grow out for what weetsie is trying to do. could you clarify on that my friend? i think i must be missing something

UPDATE:

here are a few plates i made transfers from yesterday, some of the more rhizomorphic examples of GT and AA+ cultures:




and here is a pan tropicalis (pan cyan?) that is on its 9th or 10th transfer. like i mentioned before, it  has been looking a lot whispier than other peoples' pan myc. also, it has been perfectly round on almost all of those whispy plates, showing no sectors at all, very frustrating. but i just checked my most recent transfer in that series and it looks a lot more like its supposed to. in this pic are some of the older ones and  the new one. never worked with this species before




--------------------

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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Re: Strain Isolations on Agar, Pics and Questions [Re: c10h12n2o]
    #23585370 - 08/27/16 10:22 PM (7 years, 6 months ago)

Hello guys, pardon my ignorance, but I have been wondering about this for a while and have failed to solve the problem with the search function. I see a lot of people talking about isolating cultures and making monocultures, but I have not read of any benefits of doing so. What is the benefit of creating a monoculture instead of getting a clone and leaving it at that?

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Re: Strain Isolations on Agar, Pics and Questions [Re: herrenvolk]
    #23585394 - 08/27/16 10:34 PM (7 years, 6 months ago)

It's even more consistent.  You can run actual meaningful tests on it since it is a control at that point.


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Re: Strain Isolations on Agar, Pics and Questions [Re: Inocuole]
    #23585406 - 08/27/16 10:40 PM (7 years, 6 months ago)

Are there more benfits to it? Will isolating a choice strain make for better pinsets, growth, and yields than a clone?


--------------------
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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InvisibleInocuole
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Re: Strain Isolations on Agar, Pics and Questions [Re: TedTheHighlighter]
    #23585420 - 08/27/16 10:44 PM (7 years, 6 months ago)

If that isolate does in fact have those features.  It could just as easily not.  Actually that's a misrepresentation...  It's highly unlikely that a randomly chosen isolate is going to be a better than a well chosen clone.  Isolating from a clone could produce better or worse results than the clone as it was originally, as well.


--------------------
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