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OfflineJMcDoogle
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Preserving cultures using sterile Distilled water in cryovials * 1
    #19001307 - 10/19/13 08:10 PM (10 years, 3 months ago)

Quote:

http://www.fpl.fs.fed.us/documnts/pdf1994/burds94a.pdf




This is the reference article -

Preserving Cultures Using Sterile Distiled Water in Cryovials

Abstract:

Prior to 1985, cultures at teh center for forest mycology research were maintained on a 1.5% malt extract agar test-tube slants. This system not only made it neccessary to transfer the entire collection every year but also permitted genetic change because continual growth occured. In 1985, the method of storing fungal cultures in sterile distilled water in CRYOVIALS was introduced. This study reports on the use of this method for long-term fungal storage. For varying periods of up to seven ( 7 ) years, 151 miscellaneous specials of wood-decaying Basidimycotina were stored in sterile distilled water. Water storage has numerous advantages: culture viability or growth rate is not significantly influenced; isolates can be stored longer; genetic stability is greater; the method is quick, easy, and inexpensive, and requires less space.

Cyrovials ( Internally Threaded & Self Standing

These can withstand autoclaving so if I were to buy them and try something along the lines of distilled water long term storage, I could always uses them as slighty more expensive alternative to the glass vacutainer blood collection tubes.

Heres what Im guessing - seven years, of unrefigerated storage is a possiblity, but even two years unrefigerated would be nice aswell.

Full Sterile - with option of threaded inside, or external -
which would you prefer - internal or external threads?

Take a fully colonized grain, or spore print - isolate, isolate, gather your monoculture - inside of SAB/GB/FLOW/FKN LABRATORY

Take 4-6-8 ML of Sterile Distilled Water and put onto your agar monoculture

Scrape it a bit, mix up the myc.

Suck it back up and put into the Cyrovial -

Than go about your business and save for 5 years and when needed re-enter onto agar - break down any contams if any - and sustain your strain? Or take fully colonized agar chunk, cut into 2x2 pieces and drop three or four into the cyrovial filled with sterile distilled water, seal and store. They feature a rubber type gasket internally and can be twisted completley air-tight aswell, so no need to wrap them.

Im new to all of this, so this all might be a huge waste of brainpower. Please leave any and all feedback, discreation unneccesary, so... what you think?  :sorry:


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OfflineAmanita virosa
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: JMcDoogle]
    #19026922 - 10/24/13 05:25 PM (10 years, 3 months ago)

That is an excellent article. Thanks for posting. Easy and cheap. I will try this as my culture collection has grown to the point that I don't want reslant once a year. Interesting that it seems to preserve the strain  genetically as well. That to me is almost more relevent than the longer storage time.  And he'll you could prolly just do it in micro centrifuge tubes. Smaller volume seems ideal for long term storage.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Amanita virosa]
    #19028946 - 10/24/13 11:25 PM (10 years, 3 months ago)

This is more or less the procedure we use for long term storage in my lab. 2mL cryo tubes, agar plugs, sterile water. We have a whole bloody room full of the things.


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OfflineTerry M
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: matsc]
    #19042550 - 10/27/13 08:23 PM (10 years, 3 months ago)

My experience with any kind of sterile distilled water storage showed that it doesn't work well with all species. Fortunately, it works reliably with some temperature sensitive species. Volvariella volvacea is an incredible bitch to store. It's the most finicky of all tropicals with regard to cold. I've tried slants at various temperatures, but even the highest I used, not all that much under room temperature, killed it. Fortunately, it responds very well to room temperature sterile distilled water.


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OfflineAmanita virosa
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19052808 - 10/29/13 04:37 PM (10 years, 3 months ago)

Quote:

Terry M said:
My experience with any kind of sterile distilled water storage showed that it doesn't work well with all species. Fortunately, it works reliably with some temperature sensitive species. Volvariella volvacea is an incredible bitch to store. It's the most finicky of all tropicals with regard to cold. I've tried slants at various temperatures, but even the highest I used, not all that much under room temperature, killed it. Fortunately, it responds very well to room temperature sterile distilled water.



Good to know terry. What others have you tried and how did they preserve?  I was thinking about abm and pinks.


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OfflineTerry M
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Amanita virosa]
    #19056687 - 10/30/13 09:25 AM (10 years, 3 months ago)

I don't have any specific notes on abm and pinks, but I wish I had. I was in the process of converting from slants, when I discovered that SDW was unreliable. I got frustrated and failed to make any useful records. But info on storage of these tropicals would be very good to know, as they otherwise require a special temperature. As many have found out the hard way, normal refrigerator temperatures of 40 degrees F will definitely kill them. A mini-fridge controlled to 52 degrees F works perfectly.


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Edited by Terry M (12/24/13 11:42 AM)


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OfflineTerry M
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Amanita virosa]
    #19149223 - 11/17/13 11:07 AM (10 years, 2 months ago)

Quote:

Amanita virosa said:
Quote:

Terry M said:
My experience with any kind of sterile distilled water storage showed that it doesn't work well with all species. Fortunately, it works reliably with some temperature sensitive species. Volvariella volvacea is an incredible bitch to store. It's the most finicky of all tropicals with regard to cold. I've tried slants at various temperatures, but even the highest I used, not all that much under room temperature, killed it. Fortunately, it responds very well to room temperature sterile distilled water.



Good to know terry. What others have you tried and how did they preserve?  I was thinking about abm and pinks.




Interesting news:

I was just clearing out some messy space in the basement and found a whole lot of STD vials containing assorted cultures. These are from 2011! That was when I had great hopes for SDW storage, and was buying vials and storing every culture in sight. I found some pinks, but don't think I have any ABM though I'm continuing to look. Did find some paddy straw, though.

I'm going to try and grow out a few of these 2 year old SDW cultures, but I have to be very selective as I don't want to divert a lot of time to this. If you have some other specific species you are interested in SDW storage of, let me know ASAP. I may just have it in my vial collection. Would then try to grow on a plate.

This won't be a rigorous experiment by any means, because I probably only have a couple of samples for each strain. But it may show which species it is possible to store in STD for a long time.


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OfflineJMcDoogle
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19151576 - 11/17/13 08:08 PM (10 years, 2 months ago)

:rockon:

Keep us apprised.

Glad this thread came of some use!


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: JMcDoogle]
    #19163335 - 11/20/13 01:58 AM (10 years, 2 months ago)

Personally I like to transfer about 20cc of LC to some tubes and spin them down in a rough centrifuge. Then just drain off the supernatant and replace with dH2O (3x wash/spin cycle). This removes removes the nutrient broth while allowing you to combine and concentrate your LC samples. Then just store as mentioned above.

Note. If you don't have access to a centrifuge, try positioning your tubes within your laundry during the spin cycle (never tried it but it could work).


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Tryptoman]
    #19163412 - 11/20/13 02:50 AM (10 years, 2 months ago)

at second-hand-shops you often find these hand driven salad-spin (for drying washed green-salad) bins, wich you could adapt for microcentrifugaltubes


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OfflineTerry M
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: forrest]
    #19181737 - 11/24/13 08:33 AM (10 years, 2 months ago)

Good news!

My Pink Oyster and Paddy Straw have started to grow nicely after 2 days on plates. The SDW vials were made 2 years and 1 month ago. This is particularly important for the Paddy Straw, for which I have never found a cool slant survival temperature. And I've gone as high as 58 degrees F! CORRECTION FROM 68 DEGREES

The mycelium look like the real thing, but if there are any contaminants, I'll see them in the next few days.
At some point, I will begin my "27 Year Plan." :laugh: That's storing slants with a covering of mineral oil at room temperature. This will be my emergency, archival, when all else fails culture backup. Oil covered slants are messy, so it's not for regular use. This technique was described in Perrin, Peter W, 1979 "Long term storage of wood-inhabiting fungi under mineral oil," Mycologia, 71:867-869. Perrin revived all 30 cultures after 27 years of storage. I'm planning on using PP 12x50mm culture tubes and 12 mm dual position closure caps. Can get 1,000 for only $100.


Edited by Terry M (11/24/13 09:57 AM)


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OfflineAmanita virosa
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19181780 - 11/24/13 08:52 AM (10 years, 2 months ago)

Quote:

Terry M said:
Good news!

My Pink Oyster and Paddy Straw have started to grow nicely after 2 days on plates. The SDW vials were made 2 years and 1 month ago. This is particularly important for the Paddy Straw, for which I have never found a cool slant survival temperature. And I've gone as high as 68 degrees F!

The mycelium look like the real thing, but if there are any contaminants, I'll see them in the next few days.
At some point, I will begin my "27 Year Plan." :laugh: That's storing slants with a covering of mineral oil at room temperature. This will be my emergency, archival, when all else fails culture backup. Oil covered slants are messy, so it's not for regular use. This technique was described in Perrin, Peter W, 1979 "Long term storage of wood-inhabiting fungi under mineral oil," Mycologia, 71:867-869. Perrin revived all 30 cultures after 27 years of storage. I'm planning on using PP 12x50mm culture tubes and 12 mm dual position closure caps. Can get 1,000 for only $100.



Awesome!! That's great news for all of us.


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InvisiblePestile

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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19183160 - 11/24/13 03:27 PM (10 years, 2 months ago)

Very interesting! :takingnotes:


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Offlineshopdropper
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Pestile]
    #19183497 - 11/24/13 04:44 PM (10 years, 2 months ago)

very interesting. why wood inhabiting fungi?  i wonder how well this would work for Claviceps. can you post that source paper please?


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OfflineTerry M
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: shopdropper]
    #19183632 - 11/24/13 05:19 PM (10 years, 2 months ago)

Quote:

shopdropper said:
very interesting. why wood inhabiting fungi?  i wonder how well this would work for Claviceps. can you post that source paper please?




Sorry, can't post the entire paper, as it's owned by the journal. But here's a public link to the first page, and it's only a 3-page paper.
http://www.jstor.org/discover/10.2307/3759204?uid=3739888&uid=2&uid=4&uid=3739256&sid=21102986781717


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InvisibleMrGumball
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: shopdropper]
    #19183796 - 11/24/13 05:57 PM (10 years, 2 months ago)

Quote:

shopdropper said:can you post that source paper please?




Volume 1-100 of Mycologia is available for free access at Cyberliber:  An Electronic Library for Mycology.


Edited by MrGumball (11/24/13 05:59 PM)


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: MrGumball]
    #19184926 - 11/24/13 11:06 PM (10 years, 2 months ago)

thanks.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: shopdropper]
    #19187128 - 11/25/13 01:53 PM (10 years, 2 months ago)

Im curious to see how these SDW cultures turn out. This would be a good solution for me if it does.


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Invisiblelipa

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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19200367 - 11/28/13 11:33 AM (10 years, 2 months ago)

Quote:

Terry M said:
Good news!

My Pink Oyster and Paddy Straw have started to grow nicely after 2 days on plates. The SDW vials were made 2 years and 1 month ago. This is particularly important for the Paddy Straw, for which I have never found a cool slant survival temperature. And I've gone as high as 58 degrees F! CORRECTION FROM 68 DEGREES

The mycelium look like the real thing, but if there are any contaminants, I'll see them in the next few days.
At some point, I will begin my "27 Year Plan." :laugh: That's storing slants with a covering of mineral oil at room temperature. This will be my emergency, archival, when all else fails culture backup. Oil covered slants are messy, so it's not for regular use. This technique was described in Perrin, Peter W, 1979 "Long term storage of wood-inhabiting fungi under mineral oil," Mycologia, 71:867-869. Perrin revived all 30 cultures after 27 years of storage. I'm planning on using PP 12x50mm culture tubes and 12 mm dual position closure caps. Can get 1,000 for only $100.




Works very well with Phellinus and all pleurotus. I have a good 4 years with them in sterile distilled water and I pull them every year and run them. Started 4 years ago with 10 vials each. I have not had good luck with shiitake.

Lipa


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19208250 - 11/30/13 11:31 AM (10 years, 2 months ago)

Hi Terry,

I recently recovered a culture of Volvariella volvacea that was stored at room temperature in a screw cap test tube since 23/12/2001 (almost 12 years).

The mycelium was growing nicely on agar , but unfortunately I lost it in the next day due a failure of the thermostat of the incubator, when the temperature reached 59 ° C.

The mycelium was kept in distilled water , without any piece of agar, just a big cluster of mycelium. The tubes I use has the following dimensions: 100 x 15mm diameter with 4 ml of distiled water in every tube.

The same material stored on agar and on the same date of inoculation died.


Cheers,


Wilson


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Invisiblelipa

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Re: Preserving cultures using sterile Distilled water in cryovials [Re: wbastz]
    #19216548 - 12/02/13 10:11 AM (10 years, 1 month ago)

Quote:

wbastz said:
Hi Terry,

I recently recovered a culture of Volvariella volvacea that was stored at room temperature in a screw cap test tube since 23/12/2001 (almost 12 years).

The mycelium was growing nicely on agar , but unfortunately I lost it in the next day due a failure of the thermostat of the incubator, when the temperature reached 59 ° C.

The mycelium was kept in distilled water , without any piece of agar, just a big cluster of mycelium. The tubes I use has the following dimensions: 100 x 15mm diameter with 4 ml of distiled water in every tube.

The same material stored on agar and on the same date of inoculation died.


Cheers,


Wilson




Thanks for the info. I use water agar before storage.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19298455 - 12/19/13 03:18 PM (10 years, 1 month ago)

Just coming into this.

So I can sterilize these guys filled with distilled water, I'm assuming with the lids loose, then after cool down I can either drop an agar plug into the water or place the water onto the colonized petri, "stir" it, suck the solution up and squirt into the tubes, and let sit in mid 70's room temp for years?

Does the water ever need to be oxygenated? By shaking or anything?

If not, this sounds like the storage method I've truly been waiting for...

SS


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OfflineTerry M
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Shaggy Shaman]
    #19298724 - 12/19/13 04:28 PM (10 years, 1 month ago)

Hello Shaggy,

I use small screw-top vials. I sterilize these filled with distilled water and with the caps tight, just as you would for slants. Then, under sterile conditions, I open the cap and add the culture in some form: agar plugs, loose mycellium, etc. Then I screw the cap on tightly.

You don't allow them to breath like you would slants because in the sterile distilled water there are no nutrients, so nothing to metabolize and require air for. The mycellium goes into a sort of suspended animation. If you're lucky, it will wake up after some years.

I've tried both colonized agar plugs and loose mycelium. They both work, and I don't know if one is better than the other. Either way, the cultures are a pain to fish out of the SDW. I would maybe use this method as a last resort after slants. It can't be relied on 100%, but if you mess up your slants somehow (refrigerate tropical cultures, leave the cap screwed tight and forget to loosen it and seal with parafilm, allow a contaminant to sneak in), it could save you. And storage isn't a problem because you just put them somewhere at room temperature.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Terry M]
    #19300079 - 12/19/13 09:11 PM (10 years, 1 month ago)

Thanks, Terry.

Since they aren't 100% reliable, doing multiple vials with the same genetics seems smart to me. As for fishing out the material later, why not plop the whole contents into an LC?

Picking your brain, here.


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OfflineJMcDoogle
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Shaggy Shaman]
    #19300446 - 12/19/13 10:50 PM (10 years, 1 month ago)

Sure am glad to see I've actually added something to these
forums that may after-all benefit someone!

:fuckinawesome:


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Invisiblelipa

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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Shaggy Shaman]
    #19302277 - 12/20/13 10:47 AM (10 years, 1 month ago)

Quote:

Shaggy Shaman said:
As for fishing out the material later, why not plop the whole contents into an LC?

Picking your brain, here.




One awesome way to create first generation spawn without having do a bunch of prep work every time is to do a session where you expand a lot of mycelium of a certain strain on weak nutrient agar and then throw a bunch of wedges into say 100 strait small plastic PPtubes and store. When you need to start expanding it just grab a tube and pour into sterilized grain and expand from there. They usually stay good for at least 6 month to a year before they poop out. Whats also cool is that they can be stored at room temperature.

Lipa


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19302347 - 12/20/13 11:07 AM (10 years, 1 month ago)

you mean only a wedge in a tube, without additional water?
and it won't die by lack of oxygen?


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: forrest]
    #19305690 - 12/20/13 11:31 PM (10 years, 1 month ago)

No.. there is water in the tube with the wedge.


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OfflineJMcDoogle
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19307785 - 12/21/13 12:16 PM (10 years, 1 month ago)

:smbfacepalm:


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: JMcDoogle]
    #19307967 - 12/21/13 01:02 PM (10 years, 1 month ago)

why the facepalm?

i know the thread is about storage in water, but for multiple years.

when someone gives a description of what to do but doesn't mentions to add water, and says the culture will stay good for 6 months to a year, it's not strange to think it is about something different than what the topic is about, and ask for clarification. but i guess the main difference in longivity is the low nutrient agar in stead of no nutrientss at all.

but i guess it's just stupid to ask questions, and i should go around giving people facepalms to feel better about myself.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: forrest]
    #19310151 - 12/22/13 12:55 AM (10 years, 1 month ago)

There's plenty of storage techniques out there.

Overlaying with mineral oil or adding a cryoprotectant and freezing are popular methods.

Mineral oil limits the oxygen available, thus reducing the metabolic rate.

Cryoprotectants allow you to freeze the culture, allowing you to store it essentially forever.  Milk and glycerin are popular cryoprotectants, so there's nothing fancy involved.


-FF


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I find the whole "my drug should be legal but yours should be illegal" mindset disgusting and hypocritical. It's what George Bush and company do when they drink a cocktail and debate the best way to imprison marijuana users. -Diploid


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OfflineShaggy Shaman
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: fastfred]
    #19320247 - 12/24/13 09:23 AM (10 years, 1 month ago)

Those Argos cryovials are sold out, FYI. Anyone know where else I could get some without having to spend $400?

SS


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OfflineJMcDoogle
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Shaggy Shaman]
    #19320277 - 12/24/13 09:33 AM (10 years, 1 month ago)

You could try using a variation in ML -

Here are the 2 ML Tubes.

http://www.neobits.com/argos_technologies_cry12c_internally_threaded_cryovial_p5018690.html


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: JMcDoogle]
    #19322587 - 12/24/13 06:59 PM (10 years, 1 month ago)

2 mL seems kinda small, but I'll try them out if they have them.

Thanks, guys!

SS


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19328183 - 12/26/13 10:02 AM (10 years, 1 month ago)

Quote:

lipa said:
Quote:

Shaggy Shaman said:
As for fishing out the material later, why not plop the whole contents into an LC?

Picking your brain, here.




One awesome way to create first generation spawn without having do a bunch of prep work every time is to do a session where you expand a lot of mycelium of a certain strain on weak nutrient agar and then throw a bunch of wedges into say 100 strait small plastic PPtubes and store. When you need to start expanding it just grab a tube and pour into sterilized grain and expand from there. They usually stay good for at least 6 month to a year before they poop out. Whats also cool is that they can be stored at room temperature.

Lipa




Besides the fact that they can be stored at room temp, why is this adventageous over using petri dishes?  Just make a couple, throw them in the culture fridge, whenever u need a wedge take it out, transfer, then put back into fridge.  My petris last 1-2 years in fridge.


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OfflineJMcDoogle
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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Aleon]
    #19328274 - 12/26/13 10:33 AM (10 years, 1 month ago)

Small footprint? No need for refrigeration - longer lifespan
Works with cultures that are cold sensitive, and heat sensitive, and hundreds of them can easily be stored in the same space as a normal fridge drawer.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: JMcDoogle]
    #19328278 - 12/26/13 10:35 AM (10 years, 1 month ago)

Dont need that with your red spotty one..


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Aleon]
    #19330420 - 12/26/13 08:16 PM (10 years, 1 month ago)

less work. I can make up 100 tubes pretty quick. All year I can just make a G1 jar without having to do any petri work. Yes less clutter in the lab and more room in the fridge for masters. I only do it with common varieties that I know I will be using a lot.

Lipa


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19332226 - 12/27/13 09:13 AM (10 years, 1 month ago)

Im not really sure how its less work; but if it works for you, do it.  You make less transfers and the same amount of agar using the petri method as this water method. Instead of having to go petri->vial->spawn; I just go petri->spawn; eliminating a whole step (not to mention having to sterilize your water and those 100 vials in addition to your agar.) Adding extra steps adds extra vectors for contamination.  I do 1 agar transfer to make spawn; this method uses 2.  Instead of only having 1 opportunity to contaminate via transfers, this method has 2.  Also most growers have devoted culture fridges and a stack of petri dishes shouldn't be too much to fit into one.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Aleon]
    #19334876 - 12/27/13 11:02 PM (10 years, 1 month ago)

Now I dont agree with everything yo said but, this isnt an argument, its personal preference ya know. Nobody here has said its better than anything else, we're just
trying to be informative.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Aleon]
    #19346508 - 12/30/13 02:14 PM (10 years, 1 month ago)

Quote:

Aleon said:
Im not really sure how its less work; but if it works for you, do it.  You make less transfers and the same amount of agar using the petri method as this water method. Instead of having to go petri->vial->spawn; I just go petri->spawn; eliminating a whole step (not to mention having to sterilize your water and those 100 vials in addition to your agar.) Adding extra steps adds extra vectors for contamination.  I do 1 agar transfer to make spawn; this method uses 2.  Instead of only having 1 opportunity to contaminate via transfers, this method has 2.  Also most growers have devoted culture fridges and a stack of petri dishes shouldn't be too much to fit into one.





I don't think you understand. The vials are made up the first time and only that time.  After that its vial -> grain every time for 6 months to a year.... I never get contamination doing it like this. A petri plate is more likely to contaminate in a fridge due to the fact that when you open it it is more exposed than a tiny vial. I prefer not to have to do any pre-culture work every time I have to make up a new generation.  Saves me time. I don't see how this is doing "more work".


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19346903 - 12/30/13 03:49 PM (10 years, 1 month ago)

You must make a lot of grain spawn (:

I finally figured out I only really need to make grain masters once per season, but I make a whole autoclave full of grain bags per species.


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Edited by drake89 (12/30/13 03:50 PM)


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: drake89]
    #19347902 - 12/30/13 06:50 PM (10 years, 1 month ago)

Quote:

drake89 said:
You must make a lot of grain spawn (:

I finally figured out I only really need to make grain masters once per season, but I make a whole autoclave full of grain bags per species.





Yes, I sell spawn to local farms.  I g1-g2-bulk spawn or sub.  Always going to G2 at least to keep an eye on contamination. I am not trying to push the vial thing. It just works for me and I am just saying it works period.  I do the stay at home dad thing/spawn maker/mushroom grower. Every little time I can get is precious.  It's nice to be able to sterilize and just walk in there and plop a wedge in and done. No scalpel or any other tools needed.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19350036 - 12/31/13 07:10 AM (10 years, 1 month ago)

Quote:

lipa said:
Quote:

drake89 said:
You must make a lot of grain spawn (:

I finally figured out I only really need to make grain masters once per season, but I make a whole autoclave full of grain bags per species.





Yes, I sell spawn to local farms.  I g1-g2-bulk spawn or sub.  Always going to G2 at least to keep an eye on contamination. I am not trying to push the vial thing. It just works for me and I am just saying it works period.  I do the stay at home dad thing/spawn maker/mushroom grower. Every little time I can get is precious.  It's nice to be able to sterilize and just walk in there and plop a wedge in and done. No scalpel or any other tools needed.





After you said THIS, now I see the convenience. That's nice not to have to bust out scalpel everytime u want to make masters; although I am on par with drake (I only make masters a few times a year anyways.) Also I wasn't trying to down-talk your method; I just didn't understand it fully.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: Aleon]
    #19351165 - 12/31/13 01:24 PM (10 years, 1 month ago)

Thank you for this post.  I am recovering from a 3 year hiatus from growing (had to move to a very urban spot with no room) and am in the process of trying to revive my old library that is on slants and have had no attention.  Surprisingly, there are several that are already showing some signs of life.  It looks like 50%+ are dead though.  I will be using this method (and others) into the future.


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: SnowArcher]
    #19377112 - 01/06/14 12:23 PM (10 years, 25 days ago)

would there be something against adding glycerine to agar/slant for storage in the fridge, to keep it from accidental freezing?


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: forrest]
    #19382566 - 01/07/14 01:49 PM (10 years, 23 days ago)

I'm always wary of adding glycerol to things unless its going into the deep freeze (-80C). Some fungi can eat it and keep up some slow metabolic activity we want to avoid, and as it ages it tends to form all sorts of strange little peroxides that can damage the cells. In most cases, it would be probably be safe, but why risk it.

Its also a ROYAL PAIN IN THE ASS to pipette so I prefer to avoid it whenever possible (I may be a bit bitter here after spending a week making several hundred cryo stocks of our yeast and bacterial strains in lab. I was truly a defeated man by the end of that)


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: matsc]
    #19533592 - 02/07/14 12:32 AM (9 years, 11 months ago)

thanks for that awnser matsc!

lipa, would 1,5 ml centifugetubes with myc scraped from agar, then with distilled water added so that (almost) no air is left when closing the lid, be the way to go about for the long term storage?


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: forrest]
    #19534590 - 02/07/14 09:42 AM (9 years, 11 months ago)

Personally I use 5 mil sample tubes. Strait sides. The tubes are filled to 3/4 way full of distilled water and sterilized.  I run the fungus on water agar and then drop in about 4-5 small 3-4mm squares of the samples in each tube.  Then I set them right side up in a plastic container and set on the shelve in the lab. Just this week I pulled a few out that had 2008 written on them and they have already regrown out on to the agar. They were Lentinus squarrosulus and Lentinus giganteus.

You don't cool the tubes. They sit at room temperature undisturbed. No need for anything like glycerin etc.

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Re: Preserving cultures using sterile Distilled water in cryovials [Re: lipa]
    #19535068 - 02/07/14 12:13 PM (9 years, 11 months ago)

I will be giving this a go since it seems way cheaper and more fool-proof than glass slants :thumbup:


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: drake89]
    #19538009 - 02/08/14 03:05 AM (9 years, 11 months ago)

Thanks!


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Re: Preserving cultures using sterile Distilled water in cryovials [Re: drake89]
    #19538979 - 02/08/14 10:11 AM (9 years, 11 months ago)

Quote:

drake89 said:
I will be giving this a go since it seems way cheaper and more fool-proof than glass slants :thumbup:




I would make slants and use DWS for backup. I only pull them when the slants have some issues or they get neglected in some way. Its not full-proof and there is no way to monitor them except for using them. It is a valuable tool though. Especially for cultures that have sexual and asexual lifecycles (i.e. PL. cystidiosus.)

Lipa


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