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OfflineLt.Berkenstine
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MS straight to agar for isolation: waste of time?
    #23078490 - 04/03/16 09:40 AM (8 years, 9 months ago)

So I knocked up 15 petri dishes with MS a few days ago. They look like they're doing just fine. I know that the typical practice is to clone a nice fruit to agar and then clean it up instead of straight up squirting MS onto the agar. Is this a complete waste of time? What are my chances of getting a strain that fruits poorly from isolating a very nice looking sector straight from MS?

I have a bunch of rye jars progressing very very nicely right now. Should I just toss the petris I have and start on agar when I'm done fruiting these?


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Edited by Lt.Berkenstine (04/03/16 09:43 AM)

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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine] * 1
    #23078500 - 04/03/16 09:44 AM (8 years, 9 months ago)

Nothing is guaranteed, even clones need to be tested.  But ms to agar is fine, in fact it's the best way to start because at least you can be sure of a clean culture.

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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23078542 - 04/03/16 09:58 AM (8 years, 9 months ago)

But starting from a nice clone does put odds more in my favor right? Im perfectly fine with tossing what I have and writing it off as a practice poor sesh.

I thought I remember RR talking about the inefficiency of cloning because of how many cell divisions have to take place before myc ends up being a mushroom but I'm not sure if it applies like I think it does.

Any more thoughts on cloning to agar vs MS to agar?


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Edited by Lt.Berkenstine (04/03/16 10:00 AM)

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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23078559 - 04/03/16 10:02 AM (8 years, 9 months ago)

Basically, I would hate to toss what I have and find out that cloning was completely pointless in the first place


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine] * 1
    #23078581 - 04/03/16 10:09 AM (8 years, 9 months ago)

Cloning is fine. MS is fine. You can have good results with both. Clones have the advantage of being predictable but you need to grow it at least once to know if it's good.

Sure clones have more cell divisions than a true isolate. But a well cared for clone can still be kept viable for 20 years. Yeah an isolate could be kept for longer but I will take a killer clone for 20 years over none at all.

Yes RR is essentially correct about efficiency. But that is speaking from the standpoint of a commercial grower who had been in the business for 30 years. In your case you need some fruits. So do ms, take clones, get some grows. It's all a crap shoot at first no matter what you do, so get busy if you want success.

Two of the tubs in my sig are clones. One is ms. Can you tell which ones?

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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23078602 - 04/03/16 10:18 AM (8 years, 9 months ago)

hmmm seems like the middle one has smaller fruits (or a bigger tub lol) but the one on the right looks a bit less uniform than the others. The one on the right is the MS?


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23078616 - 04/03/16 10:22 AM (8 years, 9 months ago)

Yes it is.  The tub on the left is also smaller than the others. But the point was all 3 put out a first flush of 175% BE. The number one way to achieve anything in this hobby is to get to work doing it.

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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Pastywhyte]
    #23078639 - 04/03/16 10:30 AM (8 years, 9 months ago)

can you rephrase the 175% BE part? Probably not important but I'm just curious about that haha


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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23078643 - 04/03/16 10:32 AM (8 years, 9 months ago)

Also, you're saying that the two on the left were NOT isolated, just cloned from a nice lookin specimen? Cause If that's what you're saying I think I have my answer haha


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23078654 - 04/03/16 10:35 AM (8 years, 9 months ago)

by the way I'm glad such an esteemed member of the community can take the time to help me out thanks man


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Re: MS straight to agar for isolation: waste of time? [Re: Pastywhyte]
    #23078697 - 04/03/16 10:46 AM (8 years, 9 months ago)

Ah, biological efficiency. Sweet


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23154758 - 04/25/16 06:53 PM (8 years, 8 months ago)

I'd like to bump this with a new question instead of opening a new thread. It's been 22 days since i inoculated my agar dishes with MS, and the only result so far is just a fine layer of white fuzz on the surface of the agar. It would be completely impossible to isolate a more rhizomorphic piece from what I have. One thing that I could have done wrong: I made my agar with lab grade agar from a Chem supply store near me. The directions on the package say to add more water than what I added. Agar recipes from scratch around shroomery say to add half as much as what the package said to add. So I met the directions on the lab agar and the teks on shroomery half way. I also PCed my agar with some rye jars, longer than the recommended agar cooking time.



What do you guys think fucked me up here? I have 15 plates going from B+ MS. One of them got a big old chunk of mush matter from the SS and is still just growing thin fuzzy stuff. Could it be the composition of the agar I bought? It has yeast in it...and I poorer, knocked up, and stored them to perfection. No slip ups whatsoever, and it smells like fucking sweaty feet in the bag of dishes. Is that the yeast or a terrible contamination? The final two which actually appear to have some radial chunks were kept in a separate bag and didn't smell bad at all.

Pics of the best looking dishes


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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23154765 - 04/25/16 06:54 PM (8 years, 8 months ago)

Also, no Saran Wrap was used. So I guess it wasn't stored to perfection. Just placed the dishes in a large ziplock bag blasted with Lysol.

Edited by Lt.Berkenstine (04/25/16 06:55 PM)

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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23154772 - 04/25/16 06:57 PM (8 years, 8 months ago)

first 2 pics looks like complete chaos, impossible to tell what's what in there.
you could take some transfers from 3rd and 4th pics tho and see what grows out.

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Re: MS straight to agar for isolation: waste of time? [Re: spacechildo]
    #23154855 - 04/25/16 07:20 PM (8 years, 8 months ago)

I think I'm gunna wait til I have a fruit to clone to get into agar. MS just seems hard to use for agar but then again I probably fucked up in multiple other regards. I still have about 3/4 a quart of this agar which I thinned out with water. Almost doubled its volume. If i want to start isolating I'm a couple weeks when i get some fruits, should I just get new agar?


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23154859 - 04/25/16 07:22 PM (8 years, 8 months ago)

I'm currently re- PCing the agar along with some LC jars. Anyone had experience with lab grade agar. Better yet, experience with MS to agar?


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23154866 - 04/25/16 07:24 PM (8 years, 8 months ago)

you're supposed to transfer right after you see spores germinating and myc starts to colonize the plate,
not wait until the whole plate has grown out.

just put 1 drop of spore solution in the center of the plate so you know where the myc is supposed to grow so you dont transfer from satellite contams.

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Re: MS straight to agar for isolation: waste of time? [Re: spacechildo]
    #23154926 - 04/25/16 07:45 PM (8 years, 8 months ago)

Yeah it's kind hard to get one drop on there...as far as the second part....it looks like most people wait til a while after the first signs of myc lol https://www.shroomery.org/forums/showflat.php/Number/18430998#18430998


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23154962 - 04/25/16 07:56 PM (8 years, 8 months ago)

stro put colonized grains to agar, and stro knows what to look for.
when you start from spores you can't really be sure what's germinated, that's why we transfer even if it looks clean.

You can still do a transfer from a random spot on the over grown plates, its just harder to know if its all cube myc and no contams riding along.

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Re: MS straight to agar for isolation: waste of time? [Re: spacechildo]
    #23155426 - 04/25/16 10:33 PM (8 years, 8 months ago)

actually stro cloned from fruits in that tek, and I'm trying to figure out where i went wrong in getting shitty plates. I have no problem with cleanliness, all of my jars colonize quickly and completely without problems like contams. I want to not only clean up my myc, but also isolate a strong sub-strain from some fat roots of myc.

Please read the actual inquiries and the purpose of my post before replying with trivial knowledge that wasn't even in question. Not trying to flame you, I really respect you, a great member of this site, for trying to help me, but the question was about how to prepare better dishes, not what to do with the trash I've made so far. I don't trust anything in those dishes enough to waste more materials on them.

I was confused about the water content of lab grade agar vs agar from scratch, and whether that could have been the culprit for my messy dishes. Also Whether the agar I got can even be used for cubensis or if I should start from scratch in the future. Also what MS to agar SHOULD look like vs the shit I ended up with. As I said I've watered down the same old agar and re-PCed it. I'm conflicted about whether or not to use it or get lme and agar and start from scratch. I believe the ingredients in the lab agar were yeast extract, dextrose or some other form of sugar, agar agar, perhaps more.


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine] * 1
    #23155490 - 04/25/16 10:56 PM (8 years, 8 months ago)

My first time MS to Agar.

This took about 5 transfers from a grain inoculated with MS.



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Re: MS straight to agar for isolation: waste of time? [Re: Doxx]
    #23155563 - 04/25/16 11:34 PM (8 years, 8 months ago)

I see. Nice work man! Looks beautiful. Hope I see a yield like that. I should have used a grain to knock up my dishes fosho. Still unsure whether to buy new agar or use what I have though


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Re: MS straight to agar for isolation: waste of time? [Re: Doxx]
    #23155572 - 04/25/16 11:37 PM (8 years, 8 months ago)

Quote:

Doxx said:
My first time MS to Agar.

This took about 5 transfers from a grain inoculated with MS.








Man. I'm very jealous. I really wish I could produce that, and I mean that with all due respect. Time and patience will lead me to this,I know it will.

I don't do this for profit or money, it's simply "cool" to me. I very much enjoy it. Only done MS syringes and pf cakes, one or two monotubs. I can't wait to learn more and grow all if mushrooms like you. That's so neat. Props :smile:


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Re: MS straight to agar for isolation: waste of time? [Re: Moabfighter]
    #23156212 - 04/26/16 07:45 AM (8 years, 8 months ago)

your dishes looks messy because you never did a transfer. in stro's pics you can even see the colonized grains right in the center of his plates.

If there is a question in there I missed just ask again, putting spores to agar is a pretty simple thing.

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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine] * 1
    #23156406 - 04/26/16 09:04 AM (8 years, 8 months ago)

But, Space was actually answering that question about the isolating. when you start with multi-spore to agar, no matter what, you are going to need to make a good deal of transfers before you will even begin to see sectors form to where you can start trying for isolation. also, to get any good isolation, you are going to need to clean up the myc like he was advising, also. because multi-spore to agar from a spore syringe is not going to be 100% clean. so as Space said, make some transfers when you start seeing growth, clean it up, then you will eventually start seeing sectoring, and be able to try making isolation. as for the water content thing, it should be about the same as if you used a recipe from scratch. and both will work, lab grade or scratch, whether it is made from malt extract or made with potato flakes.

also, about how it should look for MS, do as Space said, put a drop in the middle. you will see it grow from that spot. it probably isnt going to look super rhizo, but instead will fan out in a cricle and then as i said, after multiple transfers it will start to appear more rhizo and start to form sectoring.

Space knows what he is talking about. he wont lead you wrong.


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OfflineLogicaL ChaosM
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Re: MS straight to agar for isolation: waste of time? [Re: mupetmower]
    #23156584 - 04/26/16 10:06 AM (8 years, 8 months ago)

Quote:

mupetmower said:
But, Space was actually answering that question about the isolating. when you start with multi-spore to agar, no matter what, you are going to need to make a good deal of transfers before you will even begin to see sectors form to where you can start trying for isolation. also, to get any good isolation, you are going to need to clean up the myc like he was advising, also. because multi-spore to agar from a spore syringe is not going to be 100% clean. so as Space said, make some transfers when you start seeing growth, clean it up, then you will eventually start seeing sectoring, and be able to try making isolation. as for the water content thing, it should be about the same as if you used a recipe from scratch. and both will work, lab grade or scratch, whether it is made from malt extract or made with potato flakes.

also, about how it should look for MS, do as Space said, put a drop in the middle. you will see it grow from that spot. it probably isnt going to look super rhizo, but instead will fan out in a cricle and then as i said, after multiple transfers it will start to appear more rhizo and start to form sectoring.

Space knows what he is talking about. he wont lead you wrong.




QFR :takingnotes:

What do u guys think is better MS to fruit then clone on agar or MS to agar then isolate?

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Re: MS straight to agar for isolation: waste of time? [Re: LogicaL Chaos] * 1
    #23156601 - 04/26/16 10:12 AM (8 years, 8 months ago)

better how? fastest? most aggressive culture?
an isolate is just that, an isolate. its not automatically better or worse than MS, its just consistent.
MS is like rolling a dice. an isolate is like you always get the same outcome of the dice. no one can say if its a consistent 6 or 1 or even 3.

cleaning on agar, waiting for in vitro pin, putting that pin on agar, then to grains is what I like to do.

and to the OP if the main question was if your agar recipe is off, I'd say no, its solidified and you got germination and growth.
anywhere from 1-3% agar is good, you can probably go way higher too.

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Re: MS straight to agar for isolation: waste of time? [Re: spacechildo]
    #23156709 - 04/26/16 10:42 AM (8 years, 8 months ago)

hey logical, i went MS to brf cakes, and then am taking clone smaples from the best fruits to agar, as well and some spore prints on foil for agar later. i also inoc'd some plates with the same MS syringe, so it is being transfered and cleaned up right now. like space said, if you mean the fastest way, then prolly MS to brf cakes, then you have fruits that you can choose to clone and print, then start the agar process, or do like i did, and do it both ways.


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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: mupetmower]
    #23163640 - 04/28/16 12:19 AM (8 years, 8 months ago)

Not like this thread needs another bump, but Thank you guys so much for answering after my whilpersnapper comment there. I think I'm gunna keep the neater looking dishes just in case I don't have a nice fruit to clone from. But...

In a nutshell, I think I would say that MS straight to agar isn't the best idea. Doesn't mean much coming from me, but it's definitely the shittiest method of cleaning and iisolation in my humble opinion. I would personally knock up a jar and then put the grain on agar. I've already had much better results on dishes from doing this.. In my personal scenario, with the way I think, I like the idea of going from fruit to agar better. The only problem with this for the beginner is getting to the point of fruiting without having a 100% clean inoculant.

It seems like a viscous circle for a beginner to get clean myc sometimes, but the way I made sense of it is going grain to agar.

I'd like to see someone else's plates after their first innoc or even their first transfer from straight MS and see if it looks any better.


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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23164143 - 04/28/16 06:50 AM (8 years, 8 months ago)

I have done spores to agar from both prints and syringes (swabs too) dozens of times with great success. I'm not sure what you are talking about. Spores to agar is the best way to germinate spores.

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Re: MS straight to agar for isolation: waste of time? [Re: Pastywhyte]
    #23167038 - 04/28/16 09:27 PM (8 years, 8 months ago)

I guess I was looking for a certain type of growth and didn't get it. I did however get much nicer looking plates from colIbises grain though.

It's not like my failure to get what I was looking for proves anything I guess.

How did your growth look with MS to agar? How long did it take to see growth? The usual? Cause these bad boys, with a couple exceptions, took longer than expected


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OfflineLt.Berkenstine
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Re: MS straight to agar for isolation: waste of time? [Re: Lt.Berkenstine]
    #23167041 - 04/28/16 09:28 PM (8 years, 8 months ago)

The smell of the inside of my bag of dishes might also indicate that I goofed somewhere with my sterile procedure :/


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