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Containers and tek for distilled water culture storage.
    #14632590 -

I wish I could find more details.. 

Roadkill said distilled storage works. 

In 1994 The Center for Forest Mycology Research Forest Products Laboratory published an article showing agar culture plugs getting transferred to 6ml drams and cryovials.http://www.fpl.fs.fed.us/documnts/pdf1994/burds94a.pdf 

This is where everything gets fuzzy for me.  I don't know anything about these containers. I dont know how to mimic them.  Specifically their lids.  If the the myc is in stasis what does it need to be happy? :grin:

What are the down sides to this storage tech?

I have small unthreaded test tubes...If possible I would like to use them for the water storage.



Quote:
RogerRabbit http://www.shroomery.org/forums/showflat.php/Number/11494427#11494427

said:
Give it a try.  I have a few cultures in the refrigerator on distilled water and no wood that are over five years old and still viable.
RR


How?

Edited by aris (06/18/11 12:42 PM)

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Re: Containers and tek for distilled water culture storage. [Re: aris]
    #14633115 -

Its as easy as cutting wedges and dropping them under distilled water...
The mycelium will go into stasis and become dormant, waiting for proper conditions to come back to life!

I would say, that this is why some LC's last so long under refrigeration...
Distilled water + nutes...
Once the mycelium eats the nutes and grows, your left with suspended mycelium in distilled water... :super:

I have a Gym. Spc. under distilled water atm...
Got the spores from SS a few months ago, grew it out on agar, and blended up a small piece in dh2o with the eberbach...
Just sat by my flowhood for months...
Just placed it in the fridge, i bet its still VERY viable...


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Re: Containers and tek for distilled water culture storage. [Re: aris]
    #14633150 -

aris said:
I wish I could find more details.. 
This is where everything gets fuzzy for me.  I don't know anything about these containers. I dont know how to mimic them.  Specifically their lids.  If the the myc is in stasis what does it need to be happy? :grin:


Sorry, i meant to answer your question as well...
The thing about the "cyrovials" is that they generally use silicon washers, making them virtually airtight, allowing them to be submerged in liquid nitrogen tanks(dwarfs?)...

They are not saying they used liquid nitrogen, just the containers(cryovials), for their properties...
The silicon washer keeps it airtight, meaning no evaporation of your dh2o, meaning lower maintenance... :shrug:
They were all about cutting cost in that journal, so they didnt want to have to "refill" their tubes with dh2o every so often, or transfer slants.... :super:

Cryovials are pretty cheap! :super:

*Edit*
Sorry mods for the double bump...
:postcount:


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Edited by total (06/18/11 12:56 PM)

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Re: Containers and tek for distilled water culture storage. [Re: total]
    #14633169 -

Ok thanks i will "totally" seal the tubes

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Re: Containers and tek for distilled water culture storage. [Re: aris]
    #14633289 -

I use 20 ml screw-cap scintillation vials for my distilled water storage. They work great.


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Liberté, égalité, humidité.

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14636099 -

I've heard that you don't need to refrigerate them, anyone with experience on that?

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Re: Containers and tek for distilled water culture storage. [Re: OmSource]
    #14636359 -

wow .. just pulled an all-nighter.  Many many samples stored.. YAY!

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Re: Containers and tek for distilled water culture storage. [Re: OmSource]
    #14636865 -

OmSource said:
I've heard that you don't need to refrigerate them, anyone with experience on that?


That's what I understand. I haven't been refrigerating mine. Room temperature DW cultures last years, but I'd expect refrigeration to to extend this. However one of my goals for sterile distilled water storage is to get my damn long-term cultures from taking up space in the fridge!

My own "tek" for this is still being refined. This method is very susceptible to bacterial contamination. But it's been my experience so far that if there is contamination, it appears very early on. I'm growing out my DW samples on a regular basis. I'm guessing that if a culture does not show contam. after a week or two, it never will. Unless of course you are sloppy in your sterile technique and introduce contam. when fishing out an agar culture piece from the DW.


--------------------
Liberté, égalité, humidité.

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14636904 -

Terry M said:
This method is very susceptible to bacterial contamination.


A little gentamicin sulfate in your dH2O would prob help that out... :super:
It shouldnt be more bacteria prone than working with a plate per say...
Its all about procedure...


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Edited by total (06/19/11 11:18 AM)

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Re: Containers and tek for distilled water culture storage. [Re: total]
    #14637233 -

Wow, thanks for linking to that publication. Great read.

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Re: Containers and tek for distilled water culture storage. [Re: Myxomycota]
    #14637478 -

Just some extra info. I always make backups with culture tubes and distilled water. It is important that the wedges be very small 3-4mm diameter and put 3-4 of them in each tube. You don't have to use cryo-vials either. I use transport tubes that are 5ml.

There is an advantage to making distilled water vials too.  If you make a few of them for each strain it is very easy to make grain spawn masters without having to get the mycelium running on agar first. Just make up some grain and drop the wedges strait into the grain and done.

I have been thinking about growing mycelium over small pieces of wood and dropping them in to distilled water as well. You could use cardboard most likely. I have also used those little kitty litter cardboard balls they sell at the pet store moistened and loaded in small tubes with mycelium. I recently pulled a three year old shiitake culture out of the fridge and placed a small chunk on agar. Took of like a rocket and looks really healthy. Agrocybe agertita stores really well this way too.

Don't expect distilled water storage to work every time. It doesn't with every species. That's why I do both so I will know which ones will work like that in the future (I also like the ability to just make up grain spawn masters without having to have an extra plate out).  Storage vials take up very little room. That's nice when you are limited on space
.
Lipa

Edited by lipa (06/19/11 02:10 PM)

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Re: Containers and tek for distilled water culture storage. [Re: aris]
    #14637534 -

I have many zip locks like this now.



So I rubber-banded sets of 7 tubes.  I used pint jars as tube racks.  I filled the tubes with distilled h20 and foiled over each pint jar.  PC this stuff for 35 at 15psi. Allowed pc to cool in room with flow hood.  After the contents had cooled to 85-90 I cut the rubber bands.


I started transferring about 3 wedges per isolate in duplicate (2 tubes 6 samples).  I para-filmed, labeled and then put a tiny zip lock over the top and taped closed with clear packing tape. 

From 9:30 PM to 6:30 AM... that took a wayyyyyyyyyyyyyyy longer than I thought it would.
I hope this works.

Thanks for the info Lipa! 5 mush 4 you!

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Re: Containers and tek for distilled water culture storage. [Re: total]
    #14637761 -

total said:
It shouldnt be more bacteria prone than working with a plate per say...
Its all about procedure...


Nonetheless, it seems to be. I use exactly the same sterile procedures for my plates, and contamination rate for these is zilch if the culture is good and clean. It may have to do with the transfer of the plate being slow because the agar is first cut up into tiny squares, and the xfer to vials takes time. The lengthy exposure to even heavily HEPA filtered air could be the culprit. No sterilization is 100%.


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Liberté, égalité, humidité.

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14637912 -

This discussion has got me thinking. There seem to be two different approaches to sterile distilled water storage. One is to put colonized agar into distilled water. The other is to scrape and wash off mycelium from the agar and put just this into distilled water.

I use the former method, which is lengthy and difficult because I dice the agar up into tiny (2-3mm) cubes that can easily fished out with a sterile loop. I figure it is worth the trouble because I won't have to do this again for years or ever. But has anybody ever done a comparison of these methods?

EDIT: This morning I just creates some mycelium-only DW vials for 6 species. It was very easy and much less laborious than the agar cube method. I simply syringed some sterile distilled water from a vial onto one section of a 3-section plate. Then I lightly scraped the mycelium into suspension using a sterile loop, and finally sucked it up back into the syringe and squirted this into the vial. It is essentially nutritionless LC.

Also very useful would be a list of mushroom species which have not be successfully stored in sterile DW.

Edited by Terry M (06/20/11 02:57 PM)

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14660894 -

I made esentialy the same thing using vacutainers.  I made a grain mycelium water to nock up jars and the first thing I did was half fill a vacutainer and then went to my jars.  Jars show no bacterial contam so I am fairly sure the container is pure.  I will just remove a very small amount to a plate and grow it out when I need it.  This is my first attempt so the idea is not proven but in theory it is the same thing others are doing but very sterile and easy.


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I need tropical cultures, ABM, V.v....!!!!

Well things don't always look as they are and things can be misread and mistaken for what they realy are so don't read too much into what I say since I might be mistaken myself. And remember I rarely ever give a definate answer.

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Re: Containers and tek for distilled water culture storage. [Re: trout]
    #14673798 -

trout said:
I made a grain mycelium water to nock up jars and the first thing I did was half fill a vacutainer and then went to my jars.  Jars show no bacterial contam so I am fairly sure the container is pure.  I will just remove a very small amount to a plate and grow it out when I need it.


This sounds essentially like RRs abandoned grain LC.Sad to say that I tried it once last year and failed miserably. He has since rejected the tek.

I have to say that my failure hasn't totally put me off to this tek and it is encouraging to see it pop up in this most interesting thread. Having the ability to store cultures for years without doing slants is appealing. I wonder if using grains is more prone to failure than Terry M's agarLC? Although even while saying this I realize it is all about sterile technique.

My idea is basically to do what Terry did. To make several AgarLC test tubes and grow each one out on a petri or small amount of grains to test for contams. It would be nice to be able to use test tubes with rubber stoppers that could act as self healing injection ports...though I know something would have to be done to equalize the pressure in the tube, but you would only need a tiny bit to inoculate a grain master or petri dish.

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Re: Containers and tek for distilled water culture storage. [Re: trout]
    #14673843 -

trout said:
I made essentially the same thing using vacutainers.  I made a grain mycelium water to nock up jars and the first thing I did was half fill a vacutainer and then went to my jars.  Jars show no bacterial contam so I am fairly sure the container is pure.  I will just remove a very small amount to a plate and grow it out when I need it.  This is my first attempt so the idea is not proven but in theory it is the same thing others are doing but very sterile and easy.


I wouldn't expect the GLC to keep. Too nutritious from all the little bits of grain and dissolved carbohydrates that come along with it. I do GLC to make syringes. I made a batch from 3 species yesterday, and I wish I had a use for all the GLC water. It will fill 60 10 ml syringes, but I only need 4. So all the rest went down the drain this morning. :frown:


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Liberté, égalité, humidité.

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14673867 -

Sea_Goat said:
This sounds essentially like RRs abandoned grain LC.Sad to say that I tried it once last year and failed miserably. He has since rejected the tek.


Huh? Why would he abandon it for short-term storage and grain expansion? What went wrong with your attempt, Sea_Goat?


--------------------
Liberté, égalité, humidité.

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14674048 -

I have slants of the same cultures so this will just be an experiment for now.  If you think the grain LC won't keep because of the nutes wouldn't these just get used up and then you would be left with DW and myc, time will tell.  I will try the same thing with a surface scrape from agar and DW and place in a vacutainer.  I just think they will be purfect for this method of storing cultures.


--------------------
I need tropical cultures, ABM, V.v....!!!!

Well things don't always look as they are and things can be misread and mistaken for what they realy are so don't read too much into what I say since I might be mistaken myself. And remember I rarely ever give a definate answer.

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Re: Containers and tek for distilled water culture storage. [Re: Terry M]
    #14674236 -

Terry M said:
Sea_Goat said:
This sounds essentially like RRs abandoned grain LC.Sad to say that I tried it once last year and failed miserably. He has since rejected the tek.


Huh? Why would he abandon it for short-term storage and grain expansion? What went wrong with your attempt, Sea_Goat?


Bacterial contam is what went wrong with mine. I know it was my own sterile procedure that failed though. My procedure and equipment has improved greatly since my failed attempt though.

I was saying in my last post that I prefer your method of using agar instead of grains to grow a culture for long term storage.

GLC does still have it's appeal though. I like experimenting to see what works best for me...but it is helpful to get ideas from post like yours and trouts to steer my experiments in new directions....

Thanks guys

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