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InvisibleStropharis
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Stro's Cleaning and Isolating On Agar * 145
    #18430998 - 06/17/13 07:17 AM (10 years, 7 months ago)

Stro's Cleaning and Isolating On Agar




I recommend putting all your spores and clones on agar first, by doing so, you can ensure that you are working with a clean culture before you introduce it to grains, and it will also take out the guess work of spore viability.

So once you have a culture growing on agar, you will now want to make sure that it is clean and free of contaminates. How do you get there? Well for starters you need a few things, pictured below is the majority of them.



You will need a Still Air Box to do this, if you do not have one, it is a very easy build using Stro's Still Air Box. Ideally though, given that you have sufficient time and/or money, I recommend building a laminar flow hood. However, their will still be circumstances that make a still air box the more suitable choice. If you are going to build one, Stro’sFlowhood Build may be of use to you, building it was a very rewarding process. If you do decide to use it as a reference, please share your progress with me.

Some other items needed include paper towels, isopropyl alcohol, lighter, alcohol lamp or similar, gloves, scalpel, new dishes, and parafilm or cling wrap, and your cultures of course.

To demonstrate how I clean up my cultures when they need it, I will be using a few clone cultures that I took using different levels of care so they would have different levels of contamination for the purpose of this write up. A, B, and C. 

A


B


C



I don't think I have to tell you what clone was taken carelessly, the photos speak for themselves.

When taking A, I swung my un sterilized scalpel like a battle axe dismembering the pin and watched it fall onto the substrate below, then I stabbed it while doing my best Braveheart scream and preceded to gently place it on the plate. I figured this was a fairly un-climactic ending so I shook the dish and watched the nasty little pin roll around contaminating the entire medium surface.

You would ideally want to do a transfer from a dish as soon as you see growth to avoid spreading any contaminants during the transfers. The less there is growing on your dish before the transfer the better.

So now we need to clean these up right? In order to do that, we will be transferring contaminant free, healthy mycelium growth to a fresh plate. Lets use A for an example, on a culture this nasty you might want to look at it and decide where you will be taking your transfer from before you are working inside you SAB, you will be working with a limited view.

Taking another look at A; there are limited options. Using a clock position, as people often use discussing agar sectors, the best option I see here is at about 4:30 but even this area is pretty close to bacteria which I have circled in red. Care must be taken to avoid those areas so we can do this in one transfer. In addition, it is a good idea to get a good look from the back as well, sometimes bacteria can be underneath the mycelium and then you would be transferring contaminant laden mycelium. The green area is that actual wedge that I transferred from this plate to a new one.



In B you can see that there is only a small culture of bacteria that is by coincident also at the 4:30 and that all other area are suitable for transfer.


And C, does it even need a transfer? No it doesn't, you can see it is already clean culture and was ready for grains as is.  I did it anyways to keep all my clones running together and it will also further isolate the strain. 


So lets cover the transfer now;

First the working surface is wiped down with lysol and an air sanitizer sprayed into the air, and all necessary equipment wiped down with isopropyl alcohol and placed in an appropriate location relative to your work area. The bag of fresh plates was brought inside my SAB, opened, removed plates, closed, then removed from SAB. I don't normally do this but I did it to provide for more room and to facilitate this write up.

Here you can see I am just about ready to go.


All the wrap is removed from the dishes and they are set up. The dishes closest to me are ready for transfer and the ones furthest are "on deck".

Sterilize your scalpel over your alcohol flame (outside your SAB of course) and the bring it inside. Open your new dish and cool the blade on the outer perimeter of your agar. If you look, you will notice these marks on a lot of the dishes you see photos of. Here it is highlighted in blue. The mark from cooling the scalpel.


I will let the photos do most of the talking for the transfer.





A few things to note here is that my new dish is on my left side, I do this because I do not want to be reaching over it for any reason. I am right handed and will be using my right hand to do the transfers; if you were left-handed you would want to reverse this. I also do my best to have the clock position that I want easily accessible so I am not spending any more time than necessary with top off. When I transfer it to the new dish, I do my best to only reach over the agar with the scalpel and to not have my hand over the dish at any point. Normally I would complete all transfers before I wrap and mark my dishes but for the purpose of this write up, I then wrapped the new dish.



All right, that covers that for the most part. Lets talk sectoring; I will go ahead and use the images that I have up to discuss this.

Although C is a clone, it isn't a true monoculture. Sectors aren't always easy to see but most of the time you can get a general idea of sectors by the outer perimeter of the mycelia growth. I will use a bit of imagery to show you what I mean, how to select them, and explain why it is important that you don't get too wrapped up or concerned about "sectors".

I honestly don't need to isolate C any further, it is going to produce and perform well and in fact there is a possibly that isolating it further can be counter productive. I actually only took one transfer from this culture as previously discussed as I currently can't facilitate very many projects at the moment.

Moving on, if we generalize here and at a glance establish sectors based on the outer perimeter of the culture, we get something like this. You can see the fan like colonies spreading themselves out and growing at different rates and with different characteristics, sometimes separated with less dense areas refereed to as "lines of isolation". At first glance there are roughly six sectors there.



At a closer look though, there are more than that. For an example I will use only the 11:00 to 2:00 on the same image only cropped and inverted so I don't sit here all night drawing lines.

Highlighted in pink you can see even more lines of isolation and segregated individual sectors highlighted in green. Where I said we have two, well now we can see six.



I guess my point is, that it isn't important to split hairs when sectoring, as long as you are isolated away from contaminates and you are taking small transfers from the outer edge of the healthiest, strongest, and fastest rhizomorphic mycelium. If you take a small peace from the very outer edge of the area where there is one distinct sector, when there are actually three, congrats you just snatched up the fasted monoculture in the area.

Another point I would like to make while I am in the process here. Some things outside genetics can have an effect on the characteristic of your culture, whether it be rhizomorphic (strandy) and tomentose (cottony).  Things like the nutritional value of your growth medium and temperature. These plates were transferred to a different batch of agar although the measurements and method of preparation were kept the same; using Sto's Agar Prep

500ml distilled water (1/2 liter)
10g Agar-Agar
10g Malt Extract



They were however moved to a higher temperature for colonization.  There appearances speak for themselves. The colonization times were faster and characteristics are distinctive. If I don't keep medium and conditions consistent, the mycelium tricks me up. The most important thing here is we now have three clone cultures that are clean and nearly ready for grains.

Updated with a few photos, to keep things simple I will reorganize.


Original clone cultures



Seven days after first transfer from clone culture


Eleven days after first transfer from clone culture


As you can see the cultures are looking healthy and the strains ar showing their individual characteristics in A and B and as suspected C is a true monoculture.

As a learning tool, I will go over one or two things here. When I first posted the photo that were taken seven days after transfer someone was asking me a few questions and I said;

Quote:

Stropharis said:
C is now true monoculture, yes. As for A I believe there are two strains but it is hard to tell




What made me come to the conclusion that A had two strains when I posted that photo? Again it was primarily the outer parameter of the culture, although it was young and tomentose, take a look at the 12 o'Clock and you will see a very small sector with different behavior than the rest of the uniform culture.



Now just four days later, after it has matured quite a bit and has also developed rhizomorphic hyphea, you can see the contrast in uniformity between the sector at 12 o'Clock, it has fanned out and distinguished itself even greater. I was happy to see C look so clean and that sector on A distinguish itself so much after saying that.





- Stro


Edited by Stropharis (08/19/16 04:24 PM)


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OfflineMidnight Cyclone
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 5
    #18431055 - 06/17/13 07:49 AM (10 years, 7 months ago)

Very nice write-ups man!

I just read through most or all of them. SO much good information!

Thanks for sharing the knowledge :mushroom2:


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OfflineForresterM
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Re: Stro's Cleaning and Isolating On Agar [Re: Midnight Cyclone] * 1
    #18431096 - 06/17/13 08:14 AM (10 years, 7 months ago)

Very nice write up, this will be extremely helpful (along with some of your other write-ups) as I'm just starting to work with agar.  Thanks for taking the time!


--------------------
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Have some medicinal mushrooms and want to get the most out of them?  Try this double extraction method.


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InvisibleFrankHorrigan
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Re: Stro's Cleaning and Isolating On Agar [Re: Forrester] * 2
    #18431128 - 06/17/13 08:38 AM (10 years, 7 months ago)

:congrats:

I love your write ups on agar Stro.

I've been meaning to get mine done but it's not going to happen too soon it seems.

So I'm going to point people over here til then :wink: :thumbup:


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

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You should take a look. :hehehe:


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InvisibleStropharis
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Re: Stro's Cleaning and Isolating On Agar [Re: Forrester] * 2
    #18431143 - 06/17/13 08:45 AM (10 years, 7 months ago)

Forrester, nice to see you around! I really enjoy your gourmet and medicinal work, inspirational to say the least.

Thanks Frank! It was a PITA for sure. It's really is hard to discuss without stepping on toes or surpassing the scope of purpose for the write up. Hopefully it will help a few folks who don't know where or how to start.


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Invisiblebillb
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Re: Stro's Cleaning and Isolating On Agar [Re: FrankHorrigan] * 1
    #18431147 - 06/17/13 08:46 AM (10 years, 7 months ago)

Nice write up!!!


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InvisibleTonydanzawillpay
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Re: Stro's Cleaning and Isolating On Agar [Re: billb] * 1
    #18431166 - 06/17/13 08:57 AM (10 years, 7 months ago)

Great write up!


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OfflineForresterM
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #18431210 - 06/17/13 09:21 AM (10 years, 7 months ago)

Thanks for the compliment :smile:

Quote:

Stropharis said:
Hopefully it will help a few folks who don't know where or how to start.




It certainly will, agar can be daunting to get into, and I've been putting it off forever.  Having really detailed information and pictures like these helps a ton. 

Especially going to the trouble to cause infection just for demonstration, that's awesome!  I just tried my first clone of a wild reishi a couple days ago and it will really help me on cleaning it up.


--------------------
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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Invisiblegopener
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #18431261 - 06/17/13 09:36 AM (10 years, 7 months ago)

Very very helpful!!:smile:


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Offlinespacechildo
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #18431318 - 06/17/13 09:56 AM (10 years, 7 months ago)

Great write-up, cleared up a lot of things that I couldn't figure out.
I'd be truly greatful if you could elaborate this a bit for me:

Quote:

Stropharis said:
If you take a small peace from the very outer edge of the area where there is one distinct sector, when there are actually three, congrats you just snatched up the fasted monoculture in the area.

- Stro




If there's actually 3 sectors just looking like 1 for the naked eye,
is that still a monoculture?
Or did you possibly mean that it's the best place to transfer from further?

Also, your absolute last pic of Orissa C, is that a monoculture?
If not, where would you choose to transfer from/sector next?

Sorry if I'm not making sense, I think this still is a bit confusing,
and my great breakfast probably didn't make my post as accurate as I wanted it to be

:pipesmoke:


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OfflinecronicrFacebook
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Re: Stro's Cleaning and Isolating On Agar [Re: spacechildo] * 2
    #18431461 - 06/17/13 10:47 AM (10 years, 7 months ago)

:woot:
just want in on this one:thumbup:


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InvisibleStropharis
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Re: Stro's Cleaning and Isolating On Agar [Re: spacechildo] * 3
    #18431474 - 06/17/13 10:53 AM (10 years, 7 months ago)

It is only when you narrow a culture down to a smaller number of strains that sectoring becomes visible. Usually after cloning you can make out sectors, these sectors are nothing more than individual strains (monocultures/isolates) that are disdiqushing themselves by showing different growth characteristics relative to its neighboring strain creating lines of isolation between one another. When I say;

Quote:

Stropharis said:
If you take a small peace from the very outer edge of the area where there is one distinct sector, when there are actually three, congrats you just snatched up the fasted monoculture in the area.





I don't mean that these sectors are not visible to the naked but they are only noticed if you take a closer look and start "splitting hairs" or small sectors lol. If you do not need a lot of different strains from one dish then it isn't necessary, because that small peace of mycelium that you transfer will more than likely not carry all three genetics but only one. 

I want to make sure that I answer your question.

Quote:

spacechildo said:
If there's actually 3 sectors just looking like 1 for the naked eye,
is that still a monoculture?




No, a true mono-culture, isolate, or strain, should not show sectoring. When working with clones tissue on agar you have already narrowed it down and will most likely see sectors on the first culture, and although they are not true monoculture, the desired intent is met, a performing culture, therefore the reasoning behind reducing it further are limited.

Quote:

spacechildo said:
Also, your absolute last pic of Orissa C, is that a monoculture?
If not, where would you choose to transfer from/sector next?




C is now true monoculture, yes.


As for A I believe there are two strains but it is hard to tell, not relevant as I would have no reason to reduce it further.

B is not an isolate.



And you can tell why by looking where it came from.


There are so many strains in this culture that it is hard to make out sectors.

I will actually be adding to this writing up using the second dish of B to demonstrate further isolation of a strain.

I hope your answer is somewhere in my rambling.

I am late for a date but I will stop by later.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #18449350 - 06/20/13 09:00 PM (10 years, 7 months ago)

I can't believe it's been three days and there aren't at least five pages of people telling you what an excellent resource this is.  :thumbup:  Thank you for this amazing pictorial write-up.


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InvisibleStropharis
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Re: Stro's Cleaning and Isolating On Agar [Re: Psilicon] * 1
    #18450207 - 06/21/13 01:00 AM (10 years, 7 months ago)

Thanks Bro! There should be some updates coming soon.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #18450349 - 06/21/13 02:06 AM (10 years, 7 months ago)

This is an amazing write up! Props to you man:thumbup:


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OfflinePussyFart
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Re: Stro's Cleaning and Isolating On Agar [Re: newbielife5] * 2
    #18450351 - 06/21/13 02:07 AM (10 years, 7 months ago)

:whathesaid:  Very nice bro....cant't wait to see your updates.


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Offlinecubenpete
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Re: Stro's Cleaning and Isolating On Agar [Re: PussyFart] * 1
    #18451645 - 06/21/13 12:05 PM (10 years, 7 months ago)

Wow! Agar 101-04. Great read.  :takingnotes:


Edited by cubenpete (06/22/13 02:06 AM)


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InvisibleStropharis
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Re: Stro's Cleaning and Isolating On Agar [Re: cubenpete] * 1
    #18451862 - 06/21/13 12:59 PM (10 years, 7 months ago)

Updated.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #18630037 - 07/29/13 04:41 PM (10 years, 5 months ago)

Awesome dude.  Thanks for this!  :cheers:


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InvisibleStropharis
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Re: Stro's Cleaning and Isolating On Agar [Re: DeadPhan] * 2
    #18632212 - 07/30/13 01:00 AM (10 years, 5 months ago)

Glad I could help.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #18632434 - 07/30/13 04:12 AM (10 years, 5 months ago)

This makes me feel better about trying agar great write up. Loved it.


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Re: Stro's Cleaning and Isolating On Agar [Re: Crispyth]
    #18691273 - 08/11/13 12:49 PM (10 years, 5 months ago)

just posting to put in my threads :smile: starting agar soonish very nice write up though


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Offlinefirst time expert
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Re: Stro's Cleaning and Isolating On Agar [Re: vaneazy]
    #18691390 - 08/11/13 01:29 PM (10 years, 5 months ago)

Im happy to see more of these threads poppin up. When I started agar I looked for something like this and couldnt find much so I was posting questions about sectors and making iso's. this would have saved alot of time...A must read for beginers with agar!!


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InvisiblebodhisattaMDiscordReddit
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Re: Stro's Cleaning and Isolating On Agar [Re: first time expert]
    #19218374 - 12/02/13 05:36 PM (10 years, 1 month ago)

Nice write up, bump for relevancy and so that it's in my threads to use as source material


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Re: Stro's Cleaning and Isolating On Agar [Re: bodhisatta]
    #19224628 - 12/03/13 10:12 PM (10 years, 1 month ago)

Oh man thanks for the time and effort you put into that. Great read, definitely helped me out as this is exactly what I've been working more on recently.

Some notes and then a ?. In the pictures used showing 7 days after first transfer you will notice for both plates A and C that the growths for the most part are mostly cottony. Looking at the 11 days after pics you can see in them the same original cottony growth which after a certain amount of growth it develops more characteristic and rhizomorphic hyphea in some places. I believe you noted in the tek that the myc has now had time to mature and actually develop rhizomorphic hyphea at this point which is exactly what i want my question focused on. Right now I am using the same 1/2 pint wide mouth jars recommended for BRF cakes as my ajar plates. This has allowed me to reuse the same containers over and over and gain enough skill working with it to hopefully just go ahead and invest in actual agar plates. That point aside...

Is it normal for myc to grow out in that way... where the initial growth comes out more fluffy and as it matures the rhizo growth is able to form and sectoring and stuff like that become much more obvious? I ask because after reading this tek it made me wonder if perhaps all my plates simply appear so cottony because there is far less surface area for the myc to grow out and mature on like it would on a larger plate. Not a single plate that I can think of growing out over the last few months has developed nearly any "obvious" rhizo growth and its made me question a lot of different things cus well... I'd like to master the art as much as possible.

Part of the issue was likely that the agar recipe I used was an old Doc_T one that I found which worked perfectly fine for growing out clones or using colonized pieces of grain but i guess was just to solid for any sort of spore germination at all. I recently made the switch over to the same 500-10-10 you mention using which has had great success with germinating spores and well im waiting to see how the actual growth decides to come out.

SO - Would you say that those 1/2 pint wide mouths offer sufficient amount of space for myc to actually grow out and mature enough to where someone could hopefully see some solid rhizo formation or am I pushing it?

I ask because Im looking into possibley changing containers and figured a little input on the potential of this even being a factor or not is real or just something ive over thought?


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Re: Stro's Cleaning and Isolating On Agar [Re: Mosey3012]
    #19226205 - 12/04/13 11:27 AM (10 years, 1 month ago)

Very useful and helpful thread, great illustrations! Thanks for posting this information.


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Re: Stro's Cleaning and Isolating On Agar [Re: budkatz]
    #19226284 - 12/04/13 11:46 AM (10 years, 1 month ago)

Banging Write up bro

:mushroom2::mushroom2::mushroom2:
:mushroom2::mushroom2::mushroom2:


Any chance you can show something so beautiful with Pans?...


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Re: Stro's Cleaning and Isolating On Agar [Re: budkatz]
    #19226287 - 12/04/13 11:47 AM (10 years, 1 month ago)

Great guide

Isolating away from bacteria can be hard sometimes I read

thats is why the sandwich method or hot pour (on top of mycellium) was invented iirc

sometimes the bacteria will ride on the mycellium and be almost impossible to isolate away from

but I have had good success with isolating away from bacteria without once

the cause of bacteria was a biopsy too....
easy methods arent always the best :-)

agar is the only way to get stable results, anyone growing edibles should do it

oatmeal agar is an easy way to get started.. mycellium loves oatmeal agar


sometimes it is easier to start all over from sporeprint if it is hard to get rid of the bacteria
that way you can be pretty sure there is no bacteria in your dish..
only mold, and mold is easy to isolate away from

isolating away from bacteria can be hard.. antibiotic agar/hot pour/sandwich teks


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Re: Stro's Cleaning and Isolating On Agar [Re: Stool_Sample] * 1
    #19237464 - 12/06/13 05:01 PM (10 years, 1 month ago)

Quote:

Mosey3012 said:
Would you say that those 1/2 pint wide mouths offer sufficient amount of space for myc to actually grow out and mature enough to where someone could hopefully see some solid rhizo formation or am I pushing it?




Although genetics play the dominant roll, there are also many other factors that determine the growth type of your culture (rhizomorphic or tomentos). Many of times I have seen cultures start out strong and fast with rhizo, but also vice versa. As long as conditions are suitable it will continue to adapt and grow, I honestly can't tell you how much these conditions affect the uniformity, appearance, or class of mycelia it displays. I will however say that regardless, you should go ahead and move on to dishes/plates, as there are many benefits to doing so. Also, when you are starting out, try to be consistent with your agar formula and temps once you get it set up, knowing how the culture respond to it will help you when working with them.

Quote:

Stool_Sample said:
Any chance you can show something so beautiful with Pans?...




This photo was taken after a sector was removed from it, it was photoshopped back in just for you. ; )



Quote:

mio said:
thats is why the sandwich method or hot pour




I've used that method quite a bit, especially when working with stuborns and wilds.


Edited by Stropharis (12/06/13 05:04 PM)


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #19237472 - 12/06/13 05:03 PM (10 years, 1 month ago)

hey stropharis! nice to see ya around again:thumbup:


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #19237484 - 12/06/13 05:06 PM (10 years, 1 month ago)

Just stopping by, I'm hoping to be back at it in March, we will see.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #19237490 - 12/06/13 05:07 PM (10 years, 1 month ago)

ok well phone first next time:shakefist:


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to the question about agar plates vs ½pint wide mouth jars for agar [Re: cronicr]
    #19238687 - 12/06/13 10:01 PM (10 years, 1 month ago)

½ pint wide mouth works good IME, got a mono culture from 2-3 transfers from a clone it seems

now if I had taken the biopsy better and not been lazy/hurrying so much etc. it might be possible in 1 transfer

plates are just much better/more enjoyable to work with I would imagine
just be sure to get vented dishes or your culture dies

the advantage of jars for agar is you can PC them with the agar in... dishes cant be if they are plastic
(you also save the expensive parafilm, but parafilm can be substituted with cellophane wrap, it is breathable too)

larger dishes arent necessarily better, they can be harder to work with I heard
some prefer 120mm, some 80mm or so

glass dishes are harder to work with than plastic often if you want to stack them like is usually done

the biggest problem with jars IMO is that they are hard to stack after transfers
if you want to do 5-6 transfers your still air box might be over-crowded very quickly

plastic petri dishes are much better if you are serious about isolating/doing agar work IMO


Edited by lessismore (12/06/13 10:11 PM)


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Re: to the question about agar plates vs ½pint wide mouth jars for agar [Re: lessismore]
    #19333586 - 12/27/13 04:39 PM (10 years, 1 month ago)

Great write up. Thank you.


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Re: Stro's Cleaning and Isolating On Agar [Re: Chimaira]
    #19333816 - 12/27/13 05:43 PM (10 years, 1 month ago)

Awesome thread man...very informative!!!  Thanks for this:thumbup:


--------------------
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Re: Stro's Cleaning and Isolating On Agar [Re: 1down5up]
    #19335069 - 12/28/13 12:05 AM (10 years, 1 month ago)

Quote:

1down5up said:
Awesome thread man...very informative!!!  Thanks for this:thumbup:



:whathesaid:

This one can get some more front page time.


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Re: Stro's Cleaning and Isolating On Agar [Re: bodhisatta]
    #19338701 - 12/28/13 08:23 PM (10 years, 1 month ago)

:asianofapproval:
This is a beautiful writeup. Thanks bro!


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Re: Stro's Cleaning and Isolating On Agar [Re: Vis0nairee]
    #19454264 - 01/21/14 06:32 PM (10 years, 8 days ago)

Very nice guide!:thumbup:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #19654252 - 03/05/14 11:29 AM (9 years, 10 months ago)

Quote:

Stropharis said:
Just stopping by, I'm hoping to be back at it in March, we will see.



it's march, i didn't forget lol! bumpity bump stropharis where ya at!


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #19654584 - 03/05/14 01:00 PM (9 years, 10 months ago)

Feel like im in with the cool kids now with this knowledge :P. Thanks great guide ill be puttin it to use soon!


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #19673668 - 03/09/14 09:40 PM (9 years, 10 months ago)

Quote:

cronicr said:
Quote:

Stropharis said:
Just stopping by, I'm hoping to be back at it in March, we will see.



it's march, i didn't forget lol! bumpity bump stropharis where ya at!



I LOL'D! I'm in the shadows Breh!  Lately I can't help myself though and fungus is on the mind.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #19673861 - 03/09/14 10:24 PM (9 years, 10 months ago)

:toast:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #19876913 - 04/21/14 06:33 AM (9 years, 9 months ago)

I know this is old but I want ithe thread, thanks man nice pics :thumbup:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20001359 - 05/17/14 10:22 AM (9 years, 8 months ago)

wow man nice write-up!
i just jumped into agar and you've explained it very well
THAnx A10T
:mushroom2::mushroom2::mushroom2::mushroom2::mushroom2:


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Re: Stro's Cleaning and Isolating On Agar [Re: cArcace-x]
    #20001543 - 05/17/14 11:26 AM (9 years, 8 months ago)

WOW!!! Very informative & thorough thread...Thank You Stro for taking the time.


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Re: Stro's Cleaning and Isolating On Agar [Re: Keyser Soze]
    #20047148 - 05/27/14 02:13 PM (9 years, 8 months ago)

Awesome write up Stro!

Would it be best to put spores, a pin, or the inside of a fruit body onto agar?  I haven't found much info on this and it's still a little confusing to me.

Thank you!


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Re: Stro's Cleaning and Isolating On Agar [Re: magicMerlin] * 1
    #20057794 - 05/29/14 07:03 PM (9 years, 7 months ago)

Quote:

magicMerlin said:
Awesome write up Stro!

Would it be best to put spores, a pin, or the inside of a fruit body onto agar?  I haven't found much info on this and it's still a little confusing to me.

Thank you!





If you already have a multispore grow going and you want to start working with clones and/or monocultures, then it would be best to transfer a live tissue culture to agar. This clone culture would be the same genetically. However, it will more than likely still carry multiple sectors and not be a true isolate, and it also more than likely won't need to be in order to preform well.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20058076 - 05/29/14 07:57 PM (9 years, 7 months ago)

Thanks, stro

:tripping:


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Re: Stro's Cleaning and Isolating On Agar [Re: rohr]
    #20066524 - 05/31/14 08:10 PM (9 years, 7 months ago)

First of all I want to say THANK YOU, nice write up and I learned that what I thought I understood was wrong. Thanks again for the knowledge.

I have a question:

Is it possible to get a non fruiting strain from and "cloned" isolate?


Edited by Knothead (05/31/14 08:24 PM)


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Re: Stro's Cleaning and Isolating On Agar [Re: Knothead]
    #20066538 - 05/31/14 08:13 PM (9 years, 7 months ago)

Quote:

Knothead said:


Is it possible to get a non fruiting strain from and isolate?



yes, this is why they need to be tested


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Re: Stro's Cleaning and Isolating On Agar [Re: Knothead]
    #20066539 - 05/31/14 08:13 PM (9 years, 7 months ago)

I hope Stro doesn't mind me jumping in here to answer that, but yes, it's entirely possible.  You should check out pussyfart's 10 PE ISOLATES thread to see what the end result of isolation by a method very similar to this one can be.


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Agar - what, why and how?  Everything a beginner needs to know.
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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #20066590 - 05/31/14 08:25 PM (9 years, 7 months ago)

I edited the question, I meant to say "cloned" isolate. Sorry!


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Re: Stro's Cleaning and Isolating On Agar [Re: Knothead]
    #20066603 - 05/31/14 08:27 PM (9 years, 7 months ago)

Do you mean an isolate from a clone or a clone of an isolate?


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Re: Stro's Cleaning and Isolating On Agar [Re: Psilicon]
    #20066610 - 05/31/14 08:30 PM (9 years, 7 months ago)

Thanks Pussyfart, I read that a while back and didn't catch they where cloned isolates. Not trying to rub yall wrong, sometimes us all farts just don't on catch everything the first time. Thanks for the feedback as always.

Another misconception that I had. Thought I understood and found out I didn't.

Sure it's never happened to you. LOL


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Re: Stro's Cleaning and Isolating On Agar [Re: Psilicon]
    #20066615 - 05/31/14 08:31 PM (9 years, 7 months ago)

I'm referring to an isolate from a clone, Pussfart provided the answer I think.


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Re: Stro's Cleaning and Isolating On Agar [Re: Psilicon]
    #20066645 - 05/31/14 08:37 PM (9 years, 7 months ago)

Sorry VDG, thanks for pointing me to the answer on Pussyfats post! Appreciate the response, confused again.


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Re: Stro's Cleaning and Isolating On Agar [Re: Knothead]
    #20066664 - 05/31/14 08:44 PM (9 years, 7 months ago)

all isolates even from clones need to be tested, it';s rare to happen but it is possible  to get a non fruiter from a clone


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #20066724 - 05/31/14 09:02 PM (9 years, 7 months ago)

Thanks Chonicr, another misconception I had, haven't experienced it, thought I understood something and found out I was wrong.

Live, pay attention and learn.

Thanks to all!


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Re: Stro's Cleaning and Isolating On Agar [Re: Knothead]
    #20137567 - 06/15/14 11:40 PM (9 years, 7 months ago)

This is exactly what I needed as I venture onward into my quest of cloning and isolation.  Bravo, very well written and easily digested.

:takingnotes:
:vineclimb:


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Re: Stro's Cleaning and Isolating On Agar [Re: Dilated]
    #20137779 - 06/16/14 12:41 AM (9 years, 7 months ago)

Quote:

Dilated said:
This is exactly what I needed as I venture onward into my quest of cloning and isolation.  Bravo, very well written and easily digested.

:takingnotes:
:vineclimb:




QFT...im pretty sure this thread alone has helped more ppl than stropharis can imagine...


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MICROBIAL HUSBANDRY!!!!

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Re: Stro's Cleaning and Isolating On Agar [Re: blindingleaf]
    #20158049 - 06/20/14 04:30 AM (9 years, 7 months ago)

Bookmarked:thumbup:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20295300 - 07/19/14 07:30 AM (9 years, 6 months ago)

Was A subjected to any further tests to try an isolate that additional strain? Or?
Thanks for the procedure explanation! You have helped this neophyte out a great deal!


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Re: Stro's Cleaning and Isolating On Agar [Re: Cl0ud.9.Warl0ck]
    #20336354 - 07/27/14 06:10 AM (9 years, 6 months ago)

Quote:

Cl0ud.9.Warl0ck said:
Was A subjected to any further tests to try an isolate that additional strain? Or?
Thanks for the procedure explanation! You have helped this neophyte out a great deal!



I honestly don't remember, unlikely,  it served as a good learning tool at the time though.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20336892 - 07/27/14 10:16 AM (9 years, 6 months ago)

Quote:

Stropharis said:
Quote:

Cl0ud.9.Warl0ck said:
Was A subjected to any further tests to try an isolate that additional strain? Or?
Thanks for the procedure explanation! You have helped this neophyte out a great deal!



I honestly don't remember, unlikely,  it served as a good learning tool at the time though.



sneaky guy logs in like a ninja and doesn't even say hi:mad2:


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I'm tired do me a favor


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #20337632 - 07/27/14 12:52 PM (9 years, 6 months ago)

strictly business :smile: by the way thanks stro, awesome write up


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Re: Stro's Cleaning and Isolating On Agar [Re: Dilated]
    #20337650 - 07/27/14 12:55 PM (9 years, 6 months ago)

and yet i dub the unforgiven:hehehe:


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #20338325 - 07/27/14 03:10 PM (9 years, 6 months ago)

Is that a TC tag I see?  Well deserved Brother, congrats!


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20338328 - 07/27/14 03:10 PM (9 years, 6 months ago)

thnx buddy! hope life is treating ya well!


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #20343396 - 07/28/14 03:48 PM (9 years, 5 months ago)

This is an absolutely great write up. 5 for you Stropharis.


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Re: Stro's Cleaning and Isolating On Agar [Re: TheEaglesGift]
    #20408393 - 08/12/14 11:29 AM (9 years, 5 months ago)

thanks a lot for the write up!  :seriousthumbsup:

i just got agar, lme, and petri dishes in the mail today!

i have a related question that i hope no one minds answering here.....

the petri dishes i got are the polystyrene one time use kind, can i make up a batch of lmea and pour them all at once, then store them till i need them?

also i got both the three section ones (i was hoping to save space while looking for sectoring) and the non divided kind (once i get closer to isolates)
has anyone used these or have any opinions on them?


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Re: Stro's Cleaning and Isolating On Agar [Re: ant707]
    #20409680 - 08/12/14 04:56 PM (9 years, 5 months ago)

Quote:

ant707 said:
can i make up a batch of lmea and pour them all at once, then store them till i need them?




You shouldn’t have any issues if they are stored airtight in the plastic sleeves that they came in, and at room temperature. As long as they don’t dry out on you, a few months or so, you should be good. I usually just do a sleeve at a time or pour as needed.

Quote:

ant707 said: i got both the three section ones (i was hoping to save space while looking for sectoring) and the non divided kind (once i get closer to isolates)has anyone used these or have any opinions on them?




I have used the sectioned plates before and I do prefer them for this type of work, very user friendly.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20410189 - 08/12/14 07:01 PM (9 years, 5 months ago)

Quote:

Stropharis said:
You shouldn’t have any issues if they are stored airtight in the plastic sleeves that they came in, and at room temperature. As long as they don’t dry out on you, a few months or so, you should be good.




how would i go about storing them "airtight" in the sleeve they came in? vacuum seal the sleeve with a foodsaver or sumthin?

or, do you mean each individual dish airtight? like parafilm or plastic wrap?


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Re: Stro's Cleaning and Isolating On Agar [Re: ant707]
    #20410250 - 08/12/14 07:17 PM (9 years, 5 months ago)

I keep the open end of the plastic folder over, tapped shut, and the remaining unused sleeve of dishes in a small clean tupperware like container.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20410445 - 08/12/14 08:02 PM (9 years, 5 months ago)

:nodofunderstanding:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20581059 - 09/17/14 07:57 PM (9 years, 4 months ago)

Amazing write up, thank you so much for all ur help stro. :thumbup:


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Re: Stro's Cleaning and Isolating On Agar [Re: jordano]
    #20582792 - 09/18/14 08:57 AM (9 years, 4 months ago)

Wished I found this since this writeup was first conceived...

Thanks Stropharia... For your big heart and brilliance.

:getstoned:


Just awesoooooooome...
Here we goooo!!!


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Re: Stro's Cleaning and Isolating On Agar [Re: pablokabute]
    #20589513 - 09/19/14 05:24 PM (9 years, 4 months ago)

:raisemyglass:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20657997 - 10/04/14 03:39 PM (9 years, 3 months ago)

This jar of Azures is like 4 months old now it has been in the fridge foil cap on always. It is alive but in need of a transfer. I know the pic is crap but its the best I can get.

Do I remove a piece from the edge or, should I remove a piece that is bluing so I can be sure it is an active isolate? Also, more info on the jar, it got one drop of MS and appeared to be a single growth point. On potatoe water agar. Thanks


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Re: Stro's Cleaning and Isolating On Agar [Re: germish]
    #20658012 - 10/04/14 03:45 PM (9 years, 3 months ago)

always try for the leading edge, the bluing is it drying out/degrading


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Re: Stro's Cleaning and Isolating On Agar [Re: germish]
    #20659208 - 10/04/14 10:03 PM (9 years, 3 months ago)

This is exactly why I leave it to nicer people like Cron to help out.  :suicide:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20659226 - 10/04/14 10:06 PM (9 years, 3 months ago)

:hi:


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #20659236 - 10/04/14 10:08 PM (9 years, 3 months ago)

Holy shit. So the shroomery isn't dead aside from cron and Queef I mean pussyfart.

Now if only hippiechick would show back up lol.


--------------------
In the land of the blind, the one eyed man is king.

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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #20936756 - 12/06/14 08:57 AM (9 years, 1 month ago)

great thread! great info


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21137305 - 01/18/15 09:34 AM (9 years, 12 days ago)

:takingnotes:


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Re: Stro's Cleaning and Isolating On Agar [Re: PurePleasure]
    #21137338 - 01/18/15 09:43 AM (9 years, 12 days ago)

Where's Stro now?


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21137373 - 01/18/15 09:51 AM (9 years, 12 days ago)

Quote:

taGyo said:
Where's Stro now?



In the sand castle tek, hehe


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Re: Stro's Cleaning and Isolating On Agar [Re: PurePleasure]
    #21139035 - 01/18/15 02:25 PM (9 years, 11 days ago)

Quote:

taGyo said:
Where's Stro now?




Still here Breh..


Quote:

PurePleasure said:
In the sand castle tek, hehe




Not sure what you mean.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139047 - 01/18/15 02:28 PM (9 years, 11 days ago)

:hehehe:nice to see ya stro, hope lifes well


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139073 - 01/18/15 02:33 PM (9 years, 11 days ago)

Quote:

Stropharis said:
Quote:

taGyo said:
Where's Stro now?




Still here Breh..


Quote:

PurePleasure said:
In the sand castle tek, hehe




Not sure what you mean.



:rockon:


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #21139090 - 01/18/15 02:37 PM (9 years, 11 days ago)

Quote:

cronicr said:
:hehehe:nice to see ya stro, hope lifes well




So so Brother, ups and downs, ins and outs. All said and done, I'm still alive and making a living. Looks like you are doing well in the community, glad to see it.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139099 - 01/18/15 02:39 PM (9 years, 11 days ago)

Yep were still kicking lol


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #21139180 - 01/18/15 02:49 PM (9 years, 11 days ago)

Stro,

Do you have any grow logs?


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21139198 - 01/18/15 02:55 PM (9 years, 11 days ago)

no active ones atm,he graces us from time to time though


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21139399 - 01/18/15 03:20 PM (9 years, 11 days ago)

Quote:

taGyo said:
Stro,

Do you have any grow logs?




I haven't managed this link collection of old grows very well but feel free to have a look around.

Stro's Grows


Edited by Stropharis (02/23/15 05:19 PM)


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139423 - 01/18/15 03:25 PM (9 years, 11 days ago)

Wow you had KS Classic?

Fuckin' nice. I'm in love with KSSS and can't get my hands on KSC.


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21139501 - 01/18/15 03:43 PM (9 years, 11 days ago)

An old one I just patched up and added to the list, one of my favorites.

Panaeolus MS


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139511 - 01/18/15 03:44 PM (9 years, 11 days ago)

Pans > Cubes?


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21139616 - 01/18/15 04:01 PM (9 years, 11 days ago)

Depends on the notation for differentiation of greatness I suppose.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139643 - 01/18/15 04:06 PM (9 years, 11 days ago)

I turned down quite a few Pan prints 'cause I had no idea what I was getting myself into. Now I really regret it and am searching. Potency wise, I've read that Pans > Cubes but I've also read it's a slightly different trip. Did you eat those fresh or dry?


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21139678 - 01/18/15 04:12 PM (9 years, 11 days ago)

:pm:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21139748 - 01/18/15 04:28 PM (9 years, 11 days ago)

You're awesome Stro,

:rockon:


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #21139753 - 01/18/15 04:29 PM (9 years, 11 days ago)

:raisemyglass:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21242065 - 02/08/15 01:30 AM (8 years, 11 months ago)

:creepylurker:


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Re: Stro's Cleaning and Isolating On Agar [Re: Lumiz]
    #21281891 - 02/16/15 09:58 AM (8 years, 11 months ago)

Awesome write up...I understand agar work a lot more after reading this well written post.


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Re: Stro's Cleaning and Isolating On Agar [Re: blazedup]
    #21309774 - 02/21/15 05:51 PM (8 years, 11 months ago)

Just read this. Great write up! I'm looking to get into agar soon, I've bookmarked this and your agar prep write up. :raisemyglass:


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Re: Stro's Cleaning and Isolating On Agar [Re: Monty514]
    #21374281 - 03/07/15 11:36 AM (8 years, 10 months ago)

I learn more here everyday...thanks Stro +5


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Re: Stro's Cleaning and Isolating On Agar [Re: Val]
    #21463787 - 03/27/15 12:18 AM (8 years, 9 months ago)

This is wonderful.  Clear and concise. 

Greatly appreciated!


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Re: Stro's Cleaning and Isolating On Agar [Re: DirdyD]
    #21464106 - 03/27/15 03:59 AM (8 years, 9 months ago)

i have these in my notes


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Re: Stro's Cleaning and Isolating On Agar [Re: blackdust]
    #21549127 - 04/14/15 10:50 PM (8 years, 9 months ago)

This will def help me on my agar adventures. thanks :super:


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Re: Stro's Cleaning and Isolating On Agar [Re: PhosCap] * 1
    #21549761 - 04/15/15 03:18 AM (8 years, 9 months ago)

Quote:

blazedup said:
Awesome write up...I understand agar work a lot more after reading this well written post.



Quote:

Monty514 said:
Just read this. Great write up! I'm looking to get into agar soon, I've bookmarked this and your agar prep write up.



Quote:

Val said:
I learn more here everyday...thanks Stro +5



Quote:

DirdyD said:
This is wonderful.  Clear and concise. 

Greatly appreciated!



Quote:

blackdust said:
i have these in my notes



Quote:

phosi said:
This will def help me on my agar adventures. thanks :super:




Glad I could help out.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21556150 - 04/16/15 04:27 PM (8 years, 9 months ago)



Got plates in today from Medco, cynmar didn't have them in stock.
500 Tri-plate (Y) dishes $74, that is 0.14 a dish but shipping killed me and it came out to almost
0.25 a dish at a total of $125.00 delivered. I think I'm set for a while. :wink:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21556159 - 04/16/15 04:32 PM (8 years, 9 months ago)

Sounds like a reasonable deal. Pouring your own plates would be much cheaper though. I paid about 10 euro's for 100 plates, agar is about 10 euro for 100 grams (enough for 5 liter), malt extract is about 5 euro's for half a kg.


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Re: Stro's Cleaning and Isolating On Agar [Re: Albertheisenhouer]
    #21556177 - 04/16/15 04:35 PM (8 years, 9 months ago)

those are empty plates :wink:

nice stro :rockon:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21556217 - 04/16/15 04:45 PM (8 years, 9 months ago)

What size are those plates?
so with each plate you can have three projects on the go. that your plan i take it?


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Re: Stro's Cleaning and Isolating On Agar [Re: PhosCap]
    #21556242 - 04/16/15 04:52 PM (8 years, 9 months ago)

100 x 15mm, I prefer the y-plates, the most efficient use out of available work space and medium isn't cheap either. These are priced really well for segmented dishes aside from the shipping cost.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21556327 - 04/16/15 05:13 PM (8 years, 9 months ago)

Quote:

Stropharis said:
100 x 15mm, I prefer the y-plates, the most efficient use out of available work space and medium isn't cheap either. These are priced really well for segmented dishes aside from the shipping cost.




:thumbup:
Im getting those y-plates when my 100 reg plates are used up


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Re: Stro's Cleaning and Isolating On Agar [Re: PhosCap]
    #21556334 - 04/16/15 05:16 PM (8 years, 9 months ago)

Not a bad idea to keep a few sleeves of those on hand either.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21558503 - 04/17/15 06:25 AM (8 years, 9 months ago)

congrats on the tag Stro! Truly deserved! :hatsoff:


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Re: Stro's Cleaning and Isolating On Agar [Re: spacechildo]
    #21558536 - 04/17/15 06:46 AM (8 years, 9 months ago)

Quote:

spacechildo said:
congrats on the tag Stro! Truly deserved! :hatsoff:




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Re: Stro's Cleaning and Isolating On Agar [Re: bodhisatta]
    #21559226 - 04/17/15 10:49 AM (8 years, 9 months ago)

suits ya well:wink:


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #21559368 - 04/17/15 11:35 AM (8 years, 9 months ago)

Awesome thread! commenting for future reference!


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #21559580 - 04/17/15 12:38 PM (8 years, 9 months ago)

Quote:

spacechildo said:
congrats on the tag Stro! Truly deserved! :hatsoff:



Quote:

cronicr said:
suits ya well;)




:douchewink:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21562561 - 04/18/15 08:18 AM (8 years, 9 months ago)

How long can a agar plate (non inoculated) sit before contaminating? :crazy:


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Re: Stro's Cleaning and Isolating On Agar [Re: blackdust]
    #21562569 - 04/18/15 08:22 AM (8 years, 9 months ago)

Quote:

blackdust said:
How long can a agar plate (non inoculated) sit before contaminating? :crazy:



Years it will dry out first. If contamination can't get in it can't get in


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Re: Stro's Cleaning and Isolating On Agar [Re: bodhisatta] * 1
    #21562578 - 04/18/15 08:29 AM (8 years, 9 months ago)

Coming from you Bod, I feel pretty safe running a few dozen plates and then inoculate throughout the month. Running my PC can be a challenge so this will help me. Thank you.

---
Yes, i own my house and the misses approves of the grows. Just dont want my babies around the PC or telling grandma. :ooo:

---
I would like to add, I took Whytes advice on getting spores to germinate on agar and it seems to be doing the trick (for my exotics). For anyone who might want to know here it is

1. Make agar softer
2. Take blade (or other tool) and flame then coll the tool in the agar plate to make a little dip. This also get the agar a little wet in the spot.
3. use inoculating loop to transfer spores to the dip


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Re: Stro's Cleaning and Isolating On Agar [Re: blackdust]
    #21562588 - 04/18/15 08:35 AM (8 years, 9 months ago)

Leave your sab set up and keep the plates in there even so you don't have to wrap unwrap


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Re: Stro's Cleaning and Isolating On Agar [Re: spacecadetCID]
    #21930867 - 07/11/15 09:09 PM (8 years, 6 months ago)

me too


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #21952665 - 07/16/15 04:13 PM (8 years, 6 months ago)

This is very good information +5 :mushroom:


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Re: Stro's Cleaning and Isolating On Agar [Re: Dr.Satan]
    #21956759 - 07/17/15 03:49 PM (8 years, 6 months ago)

:rockon: Glad I could help!


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21957799 - 07/17/15 08:14 PM (8 years, 6 months ago)

is it just me or does a true monoculture look similar to viewing a mushroom from the bottom?


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee] * 1
    #21969404 - 07/20/15 06:13 AM (8 years, 6 months ago)



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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #21993191 - 07/24/15 11:16 PM (8 years, 6 months ago)

Just reading this over again,and I realized I never formally thanked you for this excellent write up. It has definitely helped re-ignite my love for science and biology.
I wish I had some kind of question, but your write up and question answering covers everything I could think of about isolating.

Thanks :smile:


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Offlinekiksen
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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #21993773 - 07/25/15 05:44 AM (8 years, 6 months ago)

Woow, awesome tek, i cant wait to inuculate my agar dishes, thank you!


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Re: Stro's Cleaning and Isolating On Agar [Re: kiksen]
    #22046500 - 08/05/15 10:16 AM (8 years, 5 months ago)

Hey, thanks for the write up, gotta get my agar  :nyan:


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Re: Stro's Cleaning and Isolating On Agar [Re: jcop]
    #22138476 - 08/25/15 01:51 AM (8 years, 5 months ago)

:bump2:


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Re: Stro's Cleaning and Isolating On Agar *DELETED* [Re: cronicr]
    #22139511 - 08/25/15 07:47 AM (8 years, 5 months ago)

Post deleted by abductee

Reason for deletion: nmp



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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #22139828 - 08/25/15 08:58 AM (8 years, 5 months ago)

My go-to reference for the agar work ahead.

This is really top notch material and i'm more than thankful, brother.


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Re: Stro's Cleaning and Isolating On Agar [Re: DirdyD] * 1
    #22139924 - 08/25/15 09:32 AM (8 years, 5 months ago)

yeah, this is an awesome write up.


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #22140043 - 08/25/15 10:06 AM (8 years, 5 months ago)

Don't forget to rate stro,

:super:


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #22140068 - 08/25/15 10:13 AM (8 years, 5 months ago)

I'll click around, not sure how to rate people or what it does. I think I rated someone before though.


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #22140080 - 08/25/15 10:16 AM (8 years, 5 months ago)

Click on their mushrooms below their head and add rating. It's just a nice way of thanking people for helping.

Or click on their name and go to view ratings near the bottom and add a rating. It lets everyone know how you feel about that person.

Here's Stro's page:

http://www.shroomery.org/forums/viewratings.php?type=1&usernum=170157


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #22140231 - 08/25/15 10:38 AM (8 years, 5 months ago)

thanks, I clicked on his name and saw rating and left one.


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #22140263 - 08/25/15 10:44 AM (8 years, 5 months ago)

:manofapproval:


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #22140330 - 08/25/15 10:59 AM (8 years, 5 months ago)

Dig it.

Welcome to the Shroomery. Stay humble, thankful and anything is possible.


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Re: Stro's Cleaning and Isolating On Agar [Re: DirdyD] * 1
    #22141175 - 08/25/15 02:49 PM (8 years, 5 months ago)

:cultureclash:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #22141358 - 08/25/15 03:27 PM (8 years, 5 months ago)

Lol stro. Im that white guy..haha


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee] * 1
    #22141474 - 08/25/15 03:48 PM (8 years, 5 months ago)

Rate Stro 2015 Campaign is now active.

Please sign up by rating Stro. Thank you.


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #22166849 - 08/30/15 09:34 PM (8 years, 4 months ago)

A properly organized and well articulated write up. I love it.  :congrats:


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Re: Stro's Cleaning and Isolating On Agar [Re: taGyo]
    #22530408 - 11/16/15 02:24 AM (8 years, 2 months ago)

very well written. thank you


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #23335902 - 06/12/16 10:39 AM (7 years, 7 months ago)

I've read thru a lot of this, and think I'm learning and want to try this when my agar stuff comes in.

Can someone explain just briefly - when I get a good isolated dish, and want to cut out a sector and put it into a jar of grain, how do I choose the best sector, and how big should it be? Is there any reason why I wouldn't just cut out the whole dish and stick it in the jar of grain?

Should I try to inoculate several jars from one dish?

I like to use big 2qt jars of rye, but usually knock them with syringes and want to move to agar.. still quite the noob here though.


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Re: Stro's Cleaning and Isolating On Agar [Re: ColonelAngus]
    #23335922 - 06/12/16 10:47 AM (7 years, 7 months ago)

Bigger wedge is better. You could use the whole plate. Take whatever section your eyes tell you. Since you don't know performance until you test it, what you grab doesn't matter at all really.


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OfflineCaptcrunch90
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Re: Stro's Cleaning and Isolating On Agar [Re: bodhisatta]
    #23349269 - 06/15/16 11:22 PM (7 years, 7 months ago)

Quote:

Trusted Cultivator said:
Bigger wedge is better. You could use the whole plate. Take whatever section your eyes tell you. Since you don't know performance until you test it, what you grab doesn't matter at all really.




Hey, if it's a true monoculture / isolate then you could technically take any section, and they should all perform the same, correct?


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Re: Stro's Cleaning and Isolating On Agar [Re: Captcrunch90]
    #23349396 - 06/16/16 12:23 AM (7 years, 7 months ago)

If it's a single strain isolate yes


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Re: Stro's Cleaning and Isolating On Agar [Re: bodhisatta]
    #23349412 - 06/16/16 12:29 AM (7 years, 7 months ago)

Cool thanks for the reply man.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #23589763 - 08/29/16 11:38 AM (7 years, 4 months ago)

This is very useful in clearing up some things for me being new and all.

Thank You for making this make sense!


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OfflinePortabellaFella 1
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Re: Stro's Cleaning and Isolating On Agar [Re: Mushmama]
    #23593053 - 08/30/16 10:10 AM (7 years, 4 months ago)

Same here. Thanks OP! Now I need to get rid of this super nutritious PDA I have that grows nothing more than cotton balls, mix my own and wait for something to clone so I can start to isolate. Stellar rates incoming! Thanks again!

Edit: damn 50 post req. I'll rate you soon as I hit 50!!!


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Edited by PortabellaFella 1 (08/30/16 10:12 AM)


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Re: Stro's Cleaning and Isolating On Agar [Re: PortabellaFella 1]
    #23593556 - 08/30/16 12:43 PM (7 years, 4 months ago)

I can't wait to get my new grow going so I can clone and see the difference in my work.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis] * 1
    #23658891 - 09/19/16 01:57 PM (7 years, 4 months ago)

wow, that was truly the best write up I have seen so far, thanks!
Its my first time growing on agar, and I have some questions.
I'm ten days in on the agar and so far I have some white spots about a quarter in size and growing. (first time posting and for some reason can't attach the pics I uploaded)

Anyway, I was going to wait until the jars are fully colonized and drop them into some grain, but now I'm wondering if I should isolate instead. Am I right in thinking that the purpose of isolating is to get that best mushroom possible, whereas if I dropped the mycelium as it is into the grain I'll get a mixture of the good the bad and the ugly? Thanks!


Edited by Yumbum (09/19/16 02:04 PM)


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Offlinespacechildo
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Re: Stro's Cleaning and Isolating On Agar [Re: Yumbum]
    #23658945 - 09/19/16 02:16 PM (7 years, 4 months ago)

you should do some cleaning first. take a few transfers of good healthy growth.
then put agar to grain.


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Re: Stro's Cleaning and Isolating On Agar [Re: spacechildo]
    #23658957 - 09/19/16 02:18 PM (7 years, 4 months ago)

Subscribed can never find this thread for some reason, now I can:highfive:


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Re: Stro's Cleaning and Isolating On Agar [Re: mrmazdarx9]
    #23691162 - 09/29/16 04:33 PM (7 years, 3 months ago)

Learned alot from this, thanks.

B:mushroom2::mushroom2::mushroom2::mushroom2::mushroom2:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #23945551 - 12/21/16 09:31 AM (7 years, 1 month ago)

Great stuff thanks!
:takingnotes:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #24454692 - 07/03/17 02:33 PM (6 years, 6 months ago)

That 'C' culture is a thing of beauty!


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #24592365 - 08/31/17 05:59 AM (6 years, 4 months ago)

so do I understand right that to get a nice transfer I should only cut the outer part of the myc?
I always get better and better rhizo myc when transfered and all of a sudden the next transfer gets worse.
I have always cut like the yellow triangle, but should I cut like square black like in the pic??


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OfflineQayyn Eb-Ur
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Re: Stro's Cleaning and Isolating On Agar [Re: Midnight Cyclone]
    #24660719 - 09/26/17 05:26 AM (6 years, 4 months ago)

this was precisely what I needed. I gonna get serious.


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Re: Stro's Cleaning and Isolating On Agar [Re: Qayyn Eb-Ur] * 1
    #24662748 - 09/26/17 09:19 PM (6 years, 4 months ago)

Yea this write up got me going as well.


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Re: Stro's Cleaning and Isolating On Agar [Re: abductee]
    #24810165 - 11/27/17 05:15 PM (6 years, 2 months ago)

Thanks for the explanation about sectors.


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Re: Stro's Cleaning and Isolating On Agar [Re: seekinglight] * 1
    #24817459 - 11/30/17 06:14 PM (6 years, 1 month ago)

hell yeah cool i was looking for this. thankk u


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Re: Stro's Cleaning and Isolating On Agar [Re: mycoshroom1994]
    #24854684 - 12/18/17 08:30 PM (6 years, 1 month ago)

:popcorn:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #24897404 - 01/07/18 08:30 PM (6 years, 21 days ago)

Hello. Necrobump here. Getting ready to start up my agar adventure and was wondering if this is all still viable info. Seems legit. I've read a lot of stuff over and over and this seems to line up.


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Re: Stro's Cleaning and Isolating On Agar [Re: AshSoul] * 1
    #24897486 - 01/07/18 09:13 PM (6 years, 21 days ago)

This post is legit.  :thumbup:


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #24988685 - 02/12/18 12:35 PM (5 years, 11 months ago)

If you have cloned a mono culture then grow it out will the spores from the fruit be isolated at all? Or do you just have a bunch of random spores like from most other prints?


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Re: Stro's Cleaning and Isolating On Agar [Re: Chaz08]
    #24988718 - 02/12/18 12:53 PM (5 years, 11 months ago)

Random.


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Re: Stro's Cleaning and Isolating On Agar [Re: hamloaf]
    #24990717 - 02/13/18 07:02 AM (5 years, 11 months ago)

if the plate is bacterial and swampy...

is there any way to salvage the myc in there?
maybe adding something, like, some antibiotics, perhaps, to the receiving plate?


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Re: Stro's Cleaning and Isolating On Agar [Re: pablokabute]
    #24990790 - 02/13/18 08:08 AM (5 years, 11 months ago)

There needs to be a leading edge that's not encompassed by bacteria in order for transferring away to work.  If the plate is as bad as described, consider giving Josex's biopsy method a try.
https://www.shroomery.org/forums/showflat.php/Number/24806569#24806569


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Re: Stro's Cleaning and Isolating On Agar [Re: hamloaf]
    #25043779 - 03/06/18 07:17 PM (5 years, 10 months ago)

Should we cool the scalpel in the old dish? or the new dish?

New dish is guaranteed clean, but cooling it there would involve opening it twice.

Cooling in the old dish potentially contaminates the scalpel if we cool it in the wrong place.


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Re: Stro's Cleaning and Isolating On Agar [Re: DoDecaPus]
    #25043799 - 03/06/18 07:25 PM (5 years, 10 months ago)

Old dish. The heat probably kills any bad shit anyway.


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Re: Stro's Cleaning and Isolating On Agar [Re: DoDecaPus]
    #25044091 - 03/06/18 09:46 PM (5 years, 10 months ago)

Quote:

DoDecaPus said:
Should we cool the scalpel in the old dish? or the new dish?

New dish is guaranteed clean, but cooling it there would involve opening it twice.

Cooling in the old dish potentially contaminates the scalpel if we cool it in the wrong place



I cool scalpel in old dish, but have been known to cool scalpel in new dish.  New dish, old dish, doesn't really matter.  Both should be clean.


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Re: Stro's Cleaning and Isolating On Agar [Re: hamloaf]
    #25044647 - 03/07/18 07:04 AM (5 years, 10 months ago)

Anyone can help me out.. so whats the best way to store your leftover liquid LME(I bought Coopers)? thank you. I hope its ok to ask it here as Stro called for LME in his recipe..

:smile:


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

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--- https://www.shroomery.org/forums/showflat.php/Number/23378638/fpart/1/vc/1

'The second seal: “All CONTAMINATED things and events are unsatisfactory.”'



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Edited by pablokabute (03/07/18 07:07 AM)


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Re: Stro's Cleaning and Isolating On Agar [Re: pablokabute]
    #25046920 - 03/07/18 05:54 PM (5 years, 10 months ago)

Quote:

pablokabute said:
Anyone can help me out.. so whats the best way to store your leftover liquid LME(I bought Coopers)? thank you. I hope its ok to ask it here as Stro called for LME in his recipe..

:smile:




In the container from which you poured it, in the fridge? You'll have to pressure cook it again to bring it up to temperature and kill anything that might have gotten in during ues/storage.

The best policy is probably not to have any left over. After some practice, and getting your pours consistent, the goal is to make just the right amount to fill all the plates in a sterile sleave.


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Re: Stro's Cleaning and Isolating On Agar [Re: DoDecaPus]
    #25047024 - 03/07/18 06:27 PM (5 years, 10 months ago)

Oh Im sorry for not being clear on this dodecaplus... what Im asking for is what to do with the leftover, unused malt in the can (1.1L can) after getting your due 10grams for your agar recipe... I thought I was gonna get the dry form of LME but I can only get Coopers(australian brand) and I just hope it wont spoil...and lose my 10bucks!! hahaha thanks.


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

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--- https://www.shroomery.org/forums/showflat.php/Number/23378638/fpart/1/vc/1

'The second seal: “All CONTAMINATED things and events are unsatisfactory.”'



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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25081798 - 03/22/18 08:02 AM (5 years, 10 months ago)

thanks


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25172060 - 04/28/18 04:28 PM (5 years, 8 months ago)

Thanks so much for this post really informative :bow2:


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OfflineMush Hunter
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Re: Stro's Cleaning and Isolating On Agar [Re: Shroomgirly]
    #25185347 - 05/05/18 05:21 AM (5 years, 8 months ago)

Great write up!:congrats:


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OfflinePerthMagicMan
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25216061 - 05/20/18 07:56 AM (5 years, 8 months ago)

Thanks mate, built myself a SAB and having a crack at agar


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OfflinePerthMagicMan
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25240003 - 05/31/18 08:59 AM (5 years, 7 months ago)

Here is one of my first transfers, it’s multispore from a print
I’ll pick the best looking plates, hopefully isolate and put the rest in grain jars
[url=http://files.shroomery.org


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OfflineWolfenstein23
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Re: Stro's Cleaning and Isolating On Agar [Re: PerthMagicMan]
    #25261332 - 06/11/18 11:13 AM (5 years, 7 months ago)

Stro! Thank you for this invaluable wealth of information. Your time and energy are greatly appreciated.

Hamloaf! Thank you for confirming this info is still viable.


I'm glad to see a thread with good solid info from a few years ago still active (semi at least)....Gives me warm fuzzies inside :P


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Invisibleaudiosnipez
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25303104 - 07/01/18 05:48 PM (5 years, 6 months ago)

Thank you for showing the sectoring in blue. Nice man.


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Offlinedrprobablyno
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25404850 - 08/22/18 09:37 AM (5 years, 5 months ago)

Stro, thank you so much. I learned a lot from your post. This is getting exiting.


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OfflineGilfoil
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25427470 - 08/31/18 11:34 PM (5 years, 4 months ago)

5 years plus and still good as gold, thank you good sir!


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OfflineChaos4142
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Re: Stro's Cleaning and Isolating On Agar [Re: Gilfoil]
    #25717109 - 01/01/19 06:51 PM (5 years, 27 days ago)

Thank you so much for this. This is gold standard. I am just about to do my first fruit clone n this has helped tremendously. What a write up man.


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Offlineicanusethesearch
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25717226 - 01/01/19 07:35 PM (5 years, 27 days ago)

Great Stuff very educational


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Re: Stro's Cleaning and Isolating On Agar [Re: icanusethesearch]
    #25744520 - 01/14/19 08:51 PM (5 years, 14 days ago)

Thanks for this info! I'm excited! I'll be using this tek to
MS syringe to agar to clean it up>transfer to grains>


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Re: Stro's Cleaning and Isolating On Agar [Re: woofwoof]
    #25764139 - 01/23/19 08:20 PM (5 years, 5 days ago)

:takingnotes:


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Re: Stro's Cleaning and Isolating On Agar [Re: DoDecaPus]
    #25922993 - 04/08/19 11:46 AM (4 years, 9 months ago)

I am very new,into my second flush of my first batch of brf cakes and doing my third transfer on my agar dishes today while gearing up for bulk. I have read this tek half a dozen times and each time I get more out of it. Thank you


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #25926986 - 04/10/19 02:03 PM (4 years, 9 months ago)

This piece sits beside the most helpful of all online resources explaining the fundamentals of agar work. Very nicely communicated, you're a talented and valued member of this community.

Is there any chance you'd be able to help me more clearly understand the topics of mating types, clamp connections, and fruiting viability?
Or point me in the right direction for advice 😌


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Re: Stro's Cleaning and Isolating On Agar [Re: Forrester]
    #25935613 - 04/15/19 08:17 AM (4 years, 9 months ago)

Wonderful. What determines, if a culture exhibits tomentose vs. rizhomorphic growth on agar? I have a lot of tomentose growth and hardly ever see rizomorphic growth.

Or can someone point me to a description?


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OfflineOrganica
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Re: Stro's Cleaning and Isolating On Agar [Re: Dr.Ivan]
    #25950390 - 04/23/19 07:25 AM (4 years, 9 months ago)

Great write up, time for some plates. Thanks stro!


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Re: Stro's Cleaning and Isolating On Agar [Re: Organica]
    #25950516 - 04/23/19 08:57 AM (4 years, 9 months ago)

I've grown out some agar, when can I transfer to grain??? Please help.


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Offlinewinemaker
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26035101 - 06/05/19 06:33 PM (4 years, 7 months ago)

Shoutout to Psilicon for sharing this link. First cleaning and isolating post I've read and I love it! I'll probably read up more on it just for fun but this is all I need to refer back to. Thank you very much for sharing your knowledge. <3


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26061302 - 06/19/19 08:27 AM (4 years, 7 months ago)

Thank you for this.. I learned more today than I have in the last 12 weeks.. :smile:


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Re: Stro's Cleaning and Isolating On Agar [Re: Johnny51v3]
    #26076427 - 06/27/19 05:57 AM (4 years, 6 months ago)

Nice one stro!
Saving this..
Hope all is well with you


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26178628 - 09/09/19 08:55 PM (4 years, 4 months ago)

Very nice write up, I’ll be saving this


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26221605 - 10/01/19 02:26 AM (4 years, 3 months ago)

Thanks


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Re: Stro's Cleaning and Isolating On Agar [Re: Whanjohi]
    #26267470 - 10/21/19 01:04 PM (4 years, 3 months ago)

Totally filled in a ton of blanks for me! Thank you so much you are the best


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Re: Stro's Cleaning and Isolating On Agar [Re: Bigkendog35]
    #26299226 - 11/04/19 06:55 PM (4 years, 2 months ago)

:mindblown:


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OfflineJdubbleu
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Re: Stro's Cleaning and Isolating On Agar [Re: IamMatt]
    #26307760 - 11/08/19 05:05 PM (4 years, 2 months ago)

I can see clearly now! Stoked to get to work!


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26314405 - 11/11/19 06:30 PM (4 years, 2 months ago)

:jodie:


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Re: Stro's Cleaning and Isolating On Agar [Re: ShroomTomb777]
    #26318035 - 11/13/19 03:23 PM (4 years, 2 months ago)

If i'm going from a spore print to agar, should i try to isolate from the first agar dish with multiple strains? Is it more successful/quicker to cut wedges that are multi spore and see what performs best when they fruit and clone from there?


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OfflineSporePrince
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Re: Stro's Cleaning and Isolating On Agar [Re: IamMatt]
    #26330252 - 11/19/19 04:50 AM (4 years, 2 months ago)

I'm curious to learn the reasons behind the statement "bacteria is hard to isolate away from, but mold is easy."

I had assumed it was the other way around -- mold is spewing spores, which can sit invisibly atop the mycelia, while bacteria just sits slimy wherever it is. Is it because bacteria can be growing invisibly below the mycelia?

I just transferred from 3 contaminated plates, two had spots of mold, the third had no mold but several blobs of bacteria growing in locations not connected to the mycelia. I'll see in a week or two what the transfers look like.


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OfflineMr. Boof
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26357433 - 12/02/19 03:33 PM (4 years, 1 month ago)

Hello, I have a question on what is happening when you are doing the actual transfer. You show three pictures, opening on agar with the mycelium, and then the scalpel on the fresh agar. Are you actually taking any quantity of the existing mycelium or is simply the act of slicing the existing culture and the introducing that scalpel to the fresh agar enough to begin a new growth? Thank you, and if there is more explication already listed somewhere else I’m sorry that I missed it. I am just starting but am wanting to be patient and fully understand the process before anything begins. Great write up by the way it was mostly very easy to digest as someone still learning the vocabulary.


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Re: Stro's Cleaning and Isolating On Agar [Re: Mr. Boof]
    #26357992 - 12/02/19 09:15 PM (4 years, 1 month ago)

You cut a tiny little wedge


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Offlineunibrowscowl
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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #26358048 - 12/02/19 09:59 PM (4 years, 1 month ago)

Operating under the assumption that we cannot get a true isolate with hobbyist equipment, what is the benefit of slimming down the genetics?  If I clone a mushroom to agar and spawn the myc to two different tubs with the exact same conditions, are the results going to be the same? Are they going to continue to be the same if I keep transferring and using the same clone's genetics for subsequent grows?

Has anyone out there with actual lab equipment created a true isolate that produces the same exact mushroom every time?


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Re: Stro's Cleaning and Isolating On Agar [Re: unibrowscowl]
    #26358078 - 12/02/19 10:21 PM (4 years, 1 month ago)

We can easily make isolates in our sabs we have been doing it for decades


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Offlineunibrowscowl
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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #26358095 - 12/02/19 10:36 PM (4 years, 1 month ago)

I hear you.  So they do give consistent results?

Then only thing that makes me question that they are true "monocultures" is this post from bod a little while back.

Quote:

bodhisatta said:
Quote:

denger said:
Quote:

bodhisatta said:
No that would be the third way. By the time you see mycelium on a dish its long been dikaryotic if you streaked out spores




That depends on the density of the streak. If it shows single colonies with some distance between them they could be monos with some probability.



I purposely diluted spores down quite a bit then streaked them. Looked at with a microscope there's still just too many spores. And spores like to clump. If you could dilute spores and also keep them from grabbing a partner then maybe. But even using jet dry or other surface tension agents wasn't able to get spores broken apart enough to get monokaryotic growth no matter how hard I tried. I think it would take a micromanipulator scope to achieve single spore monokaryotic growth reliably

Even with hefty agitation you'll see clumps like this


Diluted and streaked


If you made a 10cc syringe with the smallest speck of spores you can see its still too many spores


Even at the hardly visible growth phase they're long dikaryotic from streak plates(not super diluted in this case)



Ive played around a bunch with germination and trying to get monokaryotic growth but i think its out of the reach of hobby equipment


Spores clump like madmen despite your best efforts to make them disperse evenly





Is it possible that "isolates" are a group of limited genetics that produce more consistent results then MS?  I still wonder if the genetics change as you transfer the master from plate to plate?  Or does it just get more and more limited as time goes on?


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Re: Stro's Cleaning and Isolating On Agar [Re: unibrowscowl]
    #26358127 - 12/02/19 10:59 PM (4 years, 1 month ago)

In order to prove we don't have isolates you would have to prove that a fruit can contain more than one set of dikaryons in a fruit body which nobody can really prove or has so far


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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #26358167 - 12/02/19 11:46 PM (4 years, 1 month ago)

@uni I think you may be confusing isolate/strain/monoculture (in the genetic sense) with monokaryon/monokaryotic strain? When people here talk of isolates, they usually mean a dikaryotic strain that was isolated by cloning, not by isolating spores. (And after cloning, maybe subsequent transfers: but, as cronicr says, we don't know if they are required or would even work, if required: at least until someone points to a paper that checks the genetics directly.)


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Offlineunibrowscowl
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Re: Stro's Cleaning and Isolating On Agar [Re: curious.psychonaut]
    #26358176 - 12/02/19 11:57 PM (4 years, 1 month ago)

Thanks curious.

That makes sense.  I guess I just need to do some more reading up on genetics and how all that works.  I just did a biopsy on a good fruit and made my first plates a couple days ago.  Super good information to have.


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OfflineMr. Boof
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Re: Stro's Cleaning and Isolating On Agar [Re: cronicr]
    #26370931 - 12/09/19 01:19 PM (4 years, 1 month ago)

Quote:

cronicr said:
You cut a tiny little wedge




And just place it on the new agar plate? Does it need to be upside down or anything or will it be able to spread through the agar on its own?


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OfflineCrackatoa
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Re: Stro's Cleaning and Isolating On Agar [Re: Mr. Boof]
    #26370963 - 12/09/19 01:38 PM (4 years, 1 month ago)

No matter how it will grow out, but faster if you place mycellium side to agar


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Offlinedionysiusnextdoor
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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26408973 - 12/31/19 08:55 AM (4 years, 29 days ago)

Great stuff, just what I needed today!


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OfflineRay Charles
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Re: Stro's Cleaning and Isolating On Agar [Re: dionysiusnextdoor]
    #26432272 - 01/13/20 09:54 PM (4 years, 15 days ago)

I found this thread to be extremely helpful. Well done Stro. Cheers!


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OfflineTIS87
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Re: Stro's Cleaning and Isolating On Agar [Re: Ray Charles]
    #26444373 - 01/21/20 07:03 AM (4 years, 8 days ago)

This thread has been enlightening. :yeahgoodadvice:


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Re: Stro's Cleaning and Isolating On Agar [Re: TIS87]
    #26444532 - 01/21/20 09:42 AM (4 years, 8 days ago)

:takingnotes:


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Re: Stro's Cleaning and Isolating On Agar [Re: staytrippy420]
    #26444743 - 01/21/20 12:04 PM (4 years, 8 days ago)

So if myc grows over a bacteria blob how safe is it to transfer from it after it’s grown over and out past the bacteria?? It looks clean but I’m worried some bacteria will be stuck to the myc from when it grew over the colony.



Guess I could just take some and experiment.


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Re: Stro's Cleaning and Isolating On Agar [Re: A.k.a]
    #26444751 - 01/21/20 12:12 PM (4 years, 8 days ago)

I wouldn’t trust it but if you got the plates and agar ready why not? :shrug:
I always assumed once it touched any contam it was game over lol


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Re: Stro's Cleaning and Isolating On Agar [Re: staytrippy420]
    #26444763 - 01/21/20 12:19 PM (4 years, 8 days ago)

That pic is a few days old too the circles now are way past the bacteria, but I’ve also assumed it was no good.

I have another plate that swab was in first and it was totally covered in bacteria. A while after pulling it out the bacteria seemed to dry up and now mycelium is sprouting from it.

Idk if bacteria dies eventually or what but even with circular satellite colonies I’ve noticed they usually change after a while, look less watery and more solid.


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Offlinetruthseeker8987
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Re: Stro's Cleaning and Isolating On Agar [Re: A.k.a]
    #26447737 - 01/23/20 07:20 AM (4 years, 6 days ago)

Thank you for the hard earned knowledge my friend thank you


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Re: Stro's Cleaning and Isolating On Agar [Re: truthseeker8987]
    #26463896 - 02/01/20 03:44 PM (3 years, 11 months ago)

Thank you for your post Stro


--------------------
I am getting back into growing.

Been experimenting with an electric pressure cooker. 11.6psi for 2 hours seems to work.


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Re: Stro's Cleaning and Isolating On Agar [Re: Disscard]
    #26494232 - 02/19/20 11:27 PM (3 years, 11 months ago)

such a good resource here


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Re: Stro's Cleaning and Isolating On Agar [Re: woofwoof]
    #26501179 - 02/24/20 09:55 AM (3 years, 10 months ago)

Great write up!!!

I have 2 questions so anyone more experienced than me please feel free to assist me :smile:)

1. Would it matter if I clone a fruit from the first flush (would the clone also have potential to be an early bird) or later flush (would a clone from a later flush have prerequisites to also be a late bloomer?

2. If the best fruit cluster desirable to clone would be on the bottom of the substrate (maybe a stupid question but here it comes) would a clone from that cluster be more prone to grow on the bottom of a substrate as well or just where the surface conditions happens to be most suitable?

Cheers!


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Re: Stro's Cleaning and Isolating On Agar [Re: Kontiki]
    #26501264 - 02/24/20 10:53 AM (3 years, 10 months ago)

IMO
1) generally clones are first flush, and of the fastest to pin and/or cluster. People have had great clones from a second flush though.
2) cluster location doesn't matter, surface conditions are reason they are on sides or bottom


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InvisibleR3dstaffy
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. [Re: Stropharis]
    #26501930 - 02/24/20 04:59 PM (3 years, 10 months ago)

.


Edited by R3dstaffy (07/23/20 05:18 PM)


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Re: Stro's Cleaning and Isolating On Agar [Re: R3dstaffy]
    #26502091 - 02/24/20 06:41 PM (3 years, 10 months ago)

So I just want to add. Sometimes bumping an old thread can be a good thing. This was my go to for agar years ago and still is.


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Re: Stro's Cleaning and Isolating On Agar [Re: Stropharis]
    #26646463 - 05/03/20 04:39 PM (3 years, 8 months ago)

Say you're getting tomentose/fuzzy myc on your first plate after inoculating from a print to agar, and also on your transfer plate. Does further isolating allow a more healthy myc to grow? Or will it remain about the same and just ensure a clean isolation? It colonizes grain just fine, but wondering how to get a more rhizomorphic plate to isolate a monoculture.  Such as the ones you have pictured.

I'm not sure if it is the spores themselves or if it just needs further isolation.


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Offlineouuwee
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Re: Stro's Cleaning and Isolating On Agar [Re: Humlo]
    #26647897 - 05/04/20 09:46 AM (3 years, 8 months ago)

Quote:

Humlo said:
Say you're getting tomentose/fuzzy myc on your first plate after inoculating from a print to agar, and also on your transfer plate. Does further isolating allow a more healthy myc to grow? Or will it remain about the same and just ensure a clean isolation? It colonizes grain just fine, but wondering how to get a more rhizomorphic plate to isolate a monoculture.  Such as the ones you have pictured.

I'm not sure if it is the spores themselves or if it just needs further isolation.




I've seen rhizo form when u transfer further starting from tomentose. Using less nutrients when making your agar helps with rhizo because it will search for food


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OfflineHumlo
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Re: Stro's Cleaning and Isolating On Agar [Re: ouuwee]
    #26650852 - 05/05/20 01:51 PM (3 years, 8 months ago)

Awesome, thanks for the info!  I read the other day about limiting nutrients and couldn't figure out why it would have that effect.  Now it makes total sense :thumbup:


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: Humlo]
    #26664301 - 05/11/20 11:27 AM (3 years, 8 months ago)

good work


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664347 - 05/11/20 11:55 AM (3 years, 8 months ago)

amazing article


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664349 - 05/11/20 11:56 AM (3 years, 8 months ago)

good luck


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664350 - 05/11/20 11:56 AM (3 years, 8 months ago)

i like that


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664353 - 05/11/20 11:59 AM (3 years, 8 months ago)

lovely writing


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664354 - 05/11/20 12:00 PM (3 years, 8 months ago)

lovely writing. good work


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664356 - 05/11/20 12:00 PM (3 years, 8 months ago)

lovely writing. good work. nicely


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664358 - 05/11/20 12:01 PM (3 years, 8 months ago)

lovely writing. good work. nicely tek


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664359 - 05/11/20 12:02 PM (3 years, 8 months ago)

lovely writing. good work. nicely tek, good luck


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664360 - 05/11/20 12:02 PM (3 years, 8 months ago)

lovely writing. good work. nicely tek, good luck . perfect


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Offlinelamama
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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664363 - 05/11/20 12:03 PM (3 years, 8 months ago)

absolutely good


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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664367 - 05/11/20 12:05 PM (3 years, 8 months ago)

best article for noobs like me


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Re: Stro's Cleaning and Isolating On Agar [Re: lamama]
    #26664371 - 05/11/20 12:06 PM (3 years, 8 months ago)

best article for noobs like me. great man


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