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cronicr



Registered: 08/07/11
Posts: 62,905
Loc: Van Isle
Last seen: 1 minute, 59 seconds
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: cronicr] 1
#29020362 - 11/11/24 10:36 AM (2 months, 4 days ago) |
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Tub was spawned yesterday
-------------------- It doesn't matter what i think of you...all that matters is clean spawn
I'm tired do me a favor
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cronicr



Registered: 08/07/11
Posts: 62,905
Loc: Van Isle
Last seen: 1 minute, 59 seconds
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: EniQma] 1
#29020363 - 11/11/24 10:39 AM (2 months, 4 days ago) |
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Quote:
EniQma said: Would using myceliated grain rather than a wedge provide too much nutrition to stall it out?
If you let the grain consolidate itoght work or you may end up with the teeniest glc we've seen lol
-------------------- It doesn't matter what i think of you...all that matters is clean spawn
I'm tired do me a favor
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EniQma
Shroomery Addict

Registered: 11/28/23
Posts: 1,858
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: cronicr] 1
#29020369 - 11/11/24 10:42 AM (2 months, 4 days ago) |
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Quote:
cronicr said:
Quote:
EniQma said: Would using myceliated grain rather than a wedge provide too much nutrition to stall it out?
If you let the grain consolidate itoght work or you may end up with the teeniest glc we've seen lol
-------------------- Do what you need to when you have to so you can do what you want to when you can.
-Me
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SeventhMushroom
Is this thing on?


Registered: 12/30/22
Posts: 60
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: EniQma] 3
#29023263 - 11/13/24 08:23 PM (2 months, 1 day ago) |
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Quote:
EniQma said: Would using myceliated grain rather than a wedge provide too much nutrition to stall it out?
I don't think nutrient content matters too much.
For me, I use 20ml glass medicine vials with rubber stoppers, rather than the smaller plastic cryovials, but they fill the same function. I also use straight up LC solution, and have successfully revived cultures >1year old so far. I also only fill them half way, ~10ml of solution each.
I use a relatively low nutrient % anyways, 0.1% LME by weight, for that nice, clear-looking solution. However, this is plenty for the cultures to thrive. After sterilizing vials with LME solution in them, I add a small bit of existing LC, an agar wedge, or even a strip of clean, inner, mushroom flesh if I'm feeling ballsy and want to skip steps! Even with the low % of LME, the cultures take off pretty quickly.
However, the cultures *do* stall eventually - not due to lack of food in solution, but due to lack of *oxygen* in the ~10ml of air in the top half of vial. I have confirmed this by opening and closing the vials under sterile airflow, and watched as they start growing again! I even did some tests with different solution-to-air ratios, and noticed they stall faster when in *more* liquid (and thus, less air).
It also makes the whole process of reviving very easy! I just pop the vial open, and dump it on a jar of grain! just like cronicr is showing! Have not had any issues with this so far!
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cronicr



Registered: 08/07/11
Posts: 62,905
Loc: Van Isle
Last seen: 1 minute, 59 seconds
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: cronicr] 3
#29035559 - 11/25/24 12:13 AM (1 month, 21 days ago) |
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Quote:
cronicr said: Tub was spawned yesterday
 Pic was 2 days ago
-------------------- It doesn't matter what i think of you...all that matters is clean spawn
I'm tired do me a favor
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Mycoplex
Sporocarp



Registered: 10/09/21
Posts: 928
Last seen: 11 days, 7 hours
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: cronicr]
#29039160 - 11/28/24 02:18 PM (1 month, 18 days ago) |
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That tub looks great cronicr, thanks for following through with all the updates!
I also agree with the general comments above related to nutrient content. I don't think the actual amount of nutrients available at the time of storage is critical (like whether the wedge is small or not), eventually all the nutrients will become depleted after long enough storage.
I think where the size of the sample comes into play is further minimizing any contamination possibility (even considering that you're starting with a clean sample), so in that regard you probably do want to use a small sample that's hanging onto the smallest amount of agar possible, enough to give the sample structure in the vial so that the sample can sink into the distilled water and stay there.
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NeitherHere
Lucid dreamer and ClusterHead


Registered: 12/25/24
Posts: 27
Loc: NorThere
Last seen: 3 days, 18 hours
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: Mycoplex] 1
#29069605 - 12/27/24 07:25 PM (19 days, 21 hours ago) |
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Whats the general tc opinion on using something like these instead of centerfuge tubes?
They're boro, listed for use with lc, twist on lids, have a silicone/ptfe ship. They should work just fine right? I'm thinkin I could biopsy inoculate them or in oculate them with .5ml EzLc/li
I'm having an absurd time with lc and have had incredibly low success rates because i keep getting heavy sediment with honey or karo 15@30m So i'm trying to switch to this and pastys EzLc unless someones got a better suggestion https://www.amazon.com/dp/B07BHBJ4WZ/ref=sspa_mw_detail_5?ie=UTF8&psc=1&sp_csd=d2lkZ2V0TmFtZT1zcF9waG9uZV9kZXRhaWw
-------------------- Every day is a gift
Lucid dreamer, Cluster head, and part time Librarian.
Edited by NeitherHere (12/27/24 07:29 PM)
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WyoMX

Registered: 07/06/15
Posts: 2,351
Loc: PNW
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: NeitherHere] 1
#29069760 - 12/27/24 10:28 PM (19 days, 18 hours ago) |
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Not a TC but I'll throw my hat in the ring.
Those look like they'd work fine for the storage this thread is about but it'd be such a small amount of LC I don't think it's worth it. Also the sediment in your LC definitely isn't causing the contams. Could be bad inoculant or rough techniques but sediment in LC is no problem. Doesn't look pretty but will work just fine.
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Psicomb
monotub with eyes



Registered: 01/13/18
Posts: 4,917
Loc: WA
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: NeitherHere] 2
#29069766 - 12/27/24 10:46 PM (19 days, 18 hours ago) |
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Quote:
NeitherHere said: Whats the general tc opinion on using something like these instead of centerfuge tubes?
They're boro, listed for use with lc, twist on lids, have a silicone/ptfe ship. They should work just fine right? I'm thinkin I could biopsy inoculate them or in oculate them with .5ml EzLc/li
I'm having an absurd time with lc and have had incredibly low success rates because i keep getting heavy sediment with honey or karo 15@30m So i'm trying to switch to this and pastys EzLc unless someones got a better suggestion https://www.amazon.com/dp/B07BHBJ4WZ/ref=sspa_mw_detail_5?ie=UTF8&psc=1&sp_csd=d2lkZ2V0TmFtZT1zcF9waG9uZV9kZXRhaWw
What you’re asking for is related to LC; completely different than what these vials on topic here are used for.
LC is not used for long term storage. If you want to get LC done the right way, follow a tek from a TC verbatim, like Pasty’s. Those glass vials you posted are 2ml, too, which is very small to cook LC in.. just a heads up. In theory it could work fine and the plastic can take autoclave temps. But master LC the regular tried and true tek way first..
And like what was said above me, your LC isn’t contaminating because of sediment… it’s either a dirty culture or poor technique, maybe LC isn’t fully sterilized, or possible filter failure on your jars. If you’re going spores to LC, fruit biopsy to LC, or some LC you bought expanded to your own, you can’t guarantee how clean it really is until you put it on agar and can see it clearly there.
— — — —
On topic of cyrovials, I just revived some 2 year old cubensis and 2.5- 3 year old piopino cultures. This tek is the best.
Infinitely easier than slants, and last so much longer. No fridge needed.
These moved across the country with me last year and held up just fine apparently! All heil King Cryovial!! I am your servant and for you I do your bidding
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When we constantly pull things apart trying to see how it works, we may end up with only an understanding of how to destroy something
- nick sand
Edited by Psicomb (12/28/24 11:23 AM)
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NeitherHere
Lucid dreamer and ClusterHead


Registered: 12/25/24
Posts: 27
Loc: NorThere
Last seen: 3 days, 18 hours
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Re: 'Cryovial Tek': Long-Term Storage of Cultures with Cryogenic Vials [Re: Psicomb] 2
#29069882 - 12/28/24 05:39 AM (19 days, 11 hours ago) |
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I'll probably just follow this tek word for word.
currently I'm using unmodified 100ml GL45 media bottles and honey or karo as my food source. some of my bottles appear totally healthy and happy and stay that way while others appear to contaminate seemingly spontaneously or as entire batches like this last batch. regardless I'm fed up. the reason I asked about autosample bottles is because while I like the unmodified vessel approach I have a lingering suspicion that the jars that have gone contaminated after extraction have done so because of accidental introducing infection during handling in some way with the rim whether it be during sample extraction or while letting the bottles breathe.
I might play around with this all but I'll stick to the cryo bottles first as written. I'll be doing a trial run here momentarily to see what happens for me.
this is going to be a huge upgrade for me. less wasted materials, longer storage times, and legitimately clear specimens vs the headaches I've been fighting lately.
thank you guys for taking the time to give me the "down boy", like a lot of other less hands on experienced cults I sometimes struggle with not trying to be an "individual" in these endeavors :P it's a compulsion I need to work on lol. big thanks to Mycoplex for sharing !
-------------------- Every day is a gift
Lucid dreamer, Cluster head, and part time Librarian.
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