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Offlinegoldcap
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Registered: 07/16/22
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reviving +15 year old spores
    #27864522 - 07/16/22 08:14 AM (2 years, 5 months ago)

Hi.

As the title has it, this is a documentation of reviving +15 year old spores. The spores were stored sandwiched between two pieces of glass, fastened by tape. Stored in darkness in room temperature.

(Started off with like 30 prints, like 3-4 of them germinated in honey LC but mold killed off the mycelium. Now an attempt with the last good print...)

Spores were introduced to LC in a home made sterile glove box. Zero contamination so far. The jars with spores have been in 26-28 degrees, darkness for a few weeks so far. No sign of germination yet.

The liquid culture used was:
100 grams of potatoes boiled in 1 liter water until soft, filtered until liquid was more clear. 2 (European) table spoons of dextrose were mixed into the liquid and then sterilized in pressure cooker for 30 minutes.

Besides these pure liquid LC jars, some of the LC was mixed with Agar (1.5 table spoons of agar per 1 liter water) and cooked for 2 minutes before sterilization. Spores scraped with sterilized knifed and "cut" into the agar. No germination on the agar either.

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OfflineLand TroutM
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Re: reviving +15 year old spores [Re: goldcap] * 1
    #27864539 - 07/16/22 08:45 AM (2 years, 5 months ago)


Edited by Land Trout (07/16/22 09:25 AM)

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: Land Trout]
    #27867134 - 07/18/22 06:29 AM (2 years, 5 months ago)

thanks. this give me great hope :laugh:

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OnlineDERRAYLD
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Re: reviving +15 year old spores [Re: goldcap]
    #27867210 - 07/18/22 08:00 AM (2 years, 5 months ago)

I've recently germinated Puerto Rican from 04 that I've been isolating further, I used a softer agar and soaked swabs I made from spore prints in sterilized water.

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: DERRAYLD]
    #27954382 - 09/17/22 10:56 AM (2 years, 3 months ago)

Ok, good tip!

I ran out of spore prints, but in a desperate attempt I took some of the liquid culture and pored it over the agar. About 1-2mm of culture now covering the agar surface.

No success. No growth, not even from contaminants ;-).

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Invisiblebakedbeings
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Re: reviving +15 year old spores [Re: goldcap] * 2
    #27954394 - 09/17/22 11:02 AM (2 years, 3 months ago)

spores to LC seems like not the best way to do this. you are eliminating your ability to separate clean growth and instead creating an all or nothing scenario, which in the case of spore LC is almost always nothing


--------------------
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Offlinegoldcap
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Re: reviving +15 year old spores [Re: bakedbeings]
    #27999886 - 10/15/22 04:59 AM (2 years, 3 months ago)

Hoooray, there is a cloud... but first:

Quote:

Bakedbeings said:
spores to LC seems like not the best way to do this....




Yes I agree. I have now studied and made some agar. That did not work either. So I tried with "forced hydration" using a syringe and rubberbands... but well, no success so far.

However, when I was making room for the agar I removed a LC jar and put in my closet (so I could procrastinate cleaning a little longer) and what do you know, today there is a cloud in the LC. :-O

While the closet is dark most of time, every once in a while there door is open and/or the light is turned on.

The temperature is like 21 Celsius.

So the same jar that stood in min 25 to max 28 Celsius (most of the time 26 Celsius) and nothing happened, when I put it in 21 Celsius suddenly things started to happen.

Maybe a coincidence? Maybe not.... the spore print is 15-16 years old, so I will have to wait like 15-16 years to repeat this experiment.

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: goldcap]
    #27999890 - 10/15/22 05:04 AM (2 years, 3 months ago)

Ok... so now I have a cloud in the LC.

I should probably remove the cloud from LC to agar right?

What is the best way to go about it?

I am thinking that maybe I could like sterilize a spoon, scoop up the cloud and throw the thing onto an agar plate! Sounds good?

Another alternative could be to try and suck it up using a syringe...

Any other ideas?

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Invisiblebakedbeings
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Re: reviving +15 year old spores [Re: goldcap]
    #28000021 - 10/15/22 08:07 AM (2 years, 3 months ago)

yes, get the myc onto agar, however you do it

i would go with water agar. or alternatively a brf puck


--------------------
Confused? Well now you can!
HHG - cheapest way to start - how i roll

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: bakedbeings]
    #28000044 - 10/15/22 08:42 AM (2 years, 3 months ago)

Thanks for the advice!

Quote:

Bakedbeings said:
yes, get the myc onto agar, however you do it

i would go with water agar. or alternatively a brf puck




I was not aware of water agar. After doing 1 minute of research, I have learned that the basic idea is to starve bacteria of nutrients (water agar is made from distilled water) while the mycelium retain some of the nutrients from the original LC (plus some of the LC on the surface of the mycelium).

This way the healthy myc can be isolated from any bacteria.

Hmm... I like only got ONE shot at this, because I have no other spore print, so I really want to do this right.

The cloud is maybe 1-1.5 cm in diameter (perfectly circular). I will give it some time to grow out and become bigger, before I take a part of it and move to agar.

I will take one part to water agar.
And another part to nutrient agar.
And then keep rest in the LC.

Edited by goldcap (10/15/22 08:43 AM)

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OfflineRusty2096
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Re: reviving +15 year old spores [Re: goldcap]
    #28000049 - 10/15/22 08:45 AM (2 years, 3 months ago)

You have many many many shots at this - not only 1 like you said. Suck up that cloud in a syringe. Squirt 1 drop per plate (you can do both some WA plates and nutes plates)


--------------------
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We don't own things - things own us.

Semi-solid liquid culture (SSLC)

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OfflineVibetyme
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Re: reviving +15 year old spores [Re: Rusty2096] * 2
    #28000150 - 10/15/22 10:13 AM (2 years, 3 months ago)

When you have 50 posts check out the marketplace for free spores. It will show up toward the bottom of the thread list.

People around here are very very generous with their work.


--------------------
Vibetyme's LAGM 2.022

Vibetyme's LAGM_23

Vibetyme's LAGM_24

                 

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Invisiblebakedbeings
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Re: reviving +15 year old spores [Re: Rusty2096]
    #28000154 - 10/15/22 10:16 AM (2 years, 3 months ago)

Quote:

Rusty2096 said:
You have many many many shots at this - not only 1 like you said. Suck up that cloud in a syringe. Squirt 1 drop per plate (you can do both some WA plates and nutes plates)



:whathesaid:


--------------------
Confused? Well now you can!
HHG - cheapest way to start - how i roll

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: Rusty2096]
    #28001416 - 10/16/22 06:18 AM (2 years, 2 months ago)

Quote:

Rusty2096 said:
You have many many many shots at this - not only 1 like you said. Suck up that cloud in a syringe. Squirt 1 drop per plate (you can do both some WA plates and nutes plates)




Ok... that sounds really really good!

I was not aware that 1 drop is enough per plate, so thanks for telling me!

Also thanks for your advice on using a syringe! I will do this create both water agar and agar with nutrients.

Pretty wild if its possible to bring back 18<= this many year old >= 15, spores. They were stored in room temperature, darkness and printed on tinfoil, which were stored in a ziplock bag.

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: bakedbeings]
    #28001418 - 10/16/22 06:23 AM (2 years, 2 months ago)

Quote:

Bakedbeings said:
:whathesaid:




Cool! I will follow ur advice!

I am curious as to why the spores germinated when I put them in 21 Celsius degrees, while nothing happened in 26.

(Hmm, I mean it could be that it is not cubensis myc growing here)

But to be perfectly accurate the jars was _first_ placed in 26 celsius and after +2 weeks moved to 21 Celsius, so maaaybe it germinated under warmer conditions and grew out in the colder.

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: goldcap]
    #28001467 - 10/16/22 07:45 AM (2 years, 2 months ago)

Ok... I followed an old tek, so I read a little about temperatures and now I realize that the temperature I was aiming for was like far to high.

21 Celsius seems to much close to ideal growing conditions.

Apparently 23 seems to be very good temperature to have, because you should be able to have that through the whole grow process. While 24 seems to be the best, the actual temperature in a colonized jar will be slightly higher because the myc produce heat... and the less warmth, the less contaminants... 23 seems to be a sweet spot if you dont ever want to change temperature.

Also I was thinking about how to optimize the germination of super old spores using a ghetto setup. I am thinking that forced hydration using pure water... wait like 2 weeks and then make a LC and put a few drops of the spore water in the LC.

Since you can create an almost endless amount of LC jars from the sporewater, there is plenty of jars that can develop myc. If there are contaminations, the myc can be easily moved to agar for isolation.

The problem I have had with applying spore water directly onto agar, is that the spores looks too dry. In LC they can all the water it can possibly need... sometimes it takes like several weeks for this old spores to germinate. I had one print that was 15 years old, put it in a LC and after 3 months there was germination (that particular grow failed during colonization because of bacteria or something).... I mean if you like rrreally need old spores to germinate (and have time) you can do this. The agar surface seems to dry out after several weeks.

Also you save some agar... when you do the LC you always end up with more LC then you need anyway, so with many jars you can make use of all the LC.

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Offlinegoldcap
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Re: reviving +15 year old spores [Re: goldcap]
    #28001470 - 10/16/22 07:48 AM (2 years, 2 months ago)

I wonder how many newbie-no-nothin-wanna-be-growers (like me) find themselves having the problem of reviving decades old spores... can not possibly be that many.

Edited by goldcap (10/16/22 07:49 AM)

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OfflineRusty2096
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Re: reviving +15 year old spores [Re: goldcap]
    #28001490 - 10/16/22 08:12 AM (2 years, 2 months ago)

Quote:

goldcap said:
I wonder how many newbie-no-nothin-wanna-be-growers (like me) find themselves having the problem of reviving decades old spores... can not possibly be that many.




1 - you

The other noobs buy grow kits :lol:


Quote:

21 Celsius seems to much close to ideal growing conditions



21 is perfect. Higher temps you were maintening (like 26) is still inside cubes myc acceptable range but the thing is, at that temp, you will favorise bacterial growth.

Spores can take up to many weeks to germinate (especially older ones). While moving to 21c was the right thing to do , I doubt that was the turning point for germination. More likely a coincidence.


--------------------
Currently looking for nothing. You guys who sent me stuff are straight up awesome!. :mushroom2:

We don't own things - things own us.

Semi-solid liquid culture (SSLC)

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OfflineSymPlayTon
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Registered: 09/03/22
Posts: 420 420 Posts!
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Last seen: 1 year, 3 months
Re: reviving +15 year old spores [Re: Vibetyme] * 1
    #28001515 - 10/16/22 08:32 AM (2 years, 2 months ago)

Quote:

Vibetyme said:
When you have 50 posts check out the marketplace for free spores. It will show up toward the bottom of the thread list.

People around here are very very generous with their work.




I am still waiting for marketplace access lol.

Still in the 90 day waiting period lol

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OfflineSymPlayTon
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Registered: 09/03/22
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Re: reviving +15 year old spores [Re: goldcap]
    #28001518 - 10/16/22 08:36 AM (2 years, 2 months ago)

Quote:

goldcap said:
I wonder how many newbie-no-nothin-wanna-be-growers (like me) find themselves having the problem of reviving decades old spores... can not possibly be that many.





I would guess not too many lol.

I would assume the process is the same.  The big difference being more patience and more finger crossing.

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