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Backing up cultures


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#1 Jeff3

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Posted 04 March 2014 - 06:45 PM

I am sure that this has been posted here somewhere before but this is my take on it.

I do love me some slants.  They store so nice and ship well.  

Anyways this is how I go about doing mine....

 

Agar recipe

 

900 mL of water

18 grams agar powder

9 grams of LME

Coffee stir sticks cut to size for 20 X 150 threaded test tubes

 

The recipe above is for 40 slants.

 

Cut the sticks to size just to fit inside of the bevel on a 20 X 150 test tube.

I usually do a bunch at once in this case on a chop saw.

 

6176-1352057833-f9df6ffee2a57fb785f60f69f31c5257.jpg

 

Throw them in a glass of water and stick them in the microwave until the water boils.  Pull them out and let them soak while you get the agar ready.

 

Add 900 mL of warm water to a sauce pan.  Mix in the agar and LME.  Make sure it is all dissolved and bring to a simmer.  Simmer while stirring for around 10 minutes.

 

Line your tubes up in a rack.

This cheap guy works great.

Test tube rack

 

Place a stick in each of the tubes and fill about 2/3 the way with agar.  I use a 30 cc syringe to measure the agar out in each tube.  I have marked it at half and use a full syringe to fill two tubes.

 

6172-1351983999-ecb13d36b76e3bcbf54846094256db60.jpg

 

Use water at first to get your mark in the right place.  Once done you are getting about 17 cc of agar per tube.

 

6414-1354404941-ffc65735aef381ed0c81ec632b684339.jpg

 

Once filled tightened the caps.  This is the method that has worked best for me.  I have yet to have one explode or boil over.

PC for 45 minutes and let them cool until you can touch the side of the pan.  It should be hot but not pull your hand away hot.

 

Lay the whole rack on its side to achieve the angle you want.  You may want to experiment a bit before hand with water to see what give you the right angle.

I use small white dishes like you would see in a sushi restaurant for soy sauce.  Two of those stacked works well for me.

 

I let them cool in front of my hood.  You could also do it in a sanitized SAB.

 

Here is the final product.

 

9027-1373461200-4d64f0f746b0237c8482ebf276334a15.jpg

 

 

 

 


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#2 hyphaenation

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Posted 04 March 2014 - 06:48 PM

Archive material. Thanks Jeff. Can always use more agar info and especially regarding putting cultures away...
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#3 esculenta

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Posted 05 March 2014 - 12:12 AM

gr8 stuff as usual



#4 Seeker2be

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Posted 05 March 2014 - 12:43 AM

Dumb question: and the wood aids to the length of life of the culture?, trains the mycelium to consume wood? adds nutrients? or? Thanks 


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#5 dial8

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Posted 05 March 2014 - 10:05 AM

Nice thread Jeff! Very informative.



#6 Jeff3

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Posted 05 March 2014 - 11:08 AM

Dumb question: and the wood aids to the length of life of the culture?, trains the mycelium to consume wood? adds nutrients? or? Thanks


This is what I have been told. The mycelium will colonize the stick and after long storage times you are unable to get a live sample from the surface of the agar you can transfer a piece of the stick and still get a live transfer. I have never actually tried that though.
Practical purposes it seems to help retain moisture and acts like rebar for the agar, holding it together, especially when shipped.
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#7 the_chosen_one

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Posted 05 March 2014 - 12:42 PM

:ohmy: look at that saw! bring that with you this summer :wink:



#8 Spooner

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Posted 05 March 2014 - 03:17 PM

How long you expect a culture to last with the wood as spare food for the myc?

#9 Jeff3

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Posted 05 March 2014 - 03:21 PM

Two years stored in the fridge. I have heard that they last longer but I haven't tried to resurrect older than two.
One thing that will kill them fairly quick is tightening the lid. Leave it loose and parafilmed.
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#10 Jeff3

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Posted 05 March 2014 - 05:49 PM

A couple of things that I forgot to add to the OP.

 

Don't whisk the agar when you are simmering.  It introduces too much air and will foam.  It makes it really hard to measure it out correctly.

 

I didn't notice but the pic of the syringe that I posted didn't have a BD syringe cap on it.  Using a cap with the tip cut off keeps the body of the syringe out of the agar and is a ton less messy.

 

And one last thing.  Wipe the tip of your syringe before injecting it into your vials.  It keeps the rim clean from the get go and you won't have to clean them before you put the lids on.

 

*if I would have caught that sooner I would have edited the OP.  Whoops.


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#11 Myc

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Posted 06 March 2014 - 09:22 AM

**Archive Material**


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#12 Myc

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Posted 06 March 2014 - 09:27 AM

Very cool tek Jeff3.

 

My long-term storage method has been to get the mycelium onto wood dowels.

Eventually - after a very long time - the cultures will dry out from prolonged storage. I've recently learned this the hard way since I neglected my reishi culture and cannot seem to revive it even after transferring dowel material.

I believe I've had this culture for around 7 years now. It has remained on the original dowels all this time and kept in the 'fridge. I went to make transfers for a fellow member and have been having heck getting the fungus to run again.

 

Your idea is much more manageable as far as handling and storage.

Extremely smart is the addition of the coffee stirrer - just brilliant!!



#13 Seeker2be

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Posted 06 March 2014 - 10:00 AM

what about making slight perforations in the stick not so it will break but to allow for multiple stick piece transfers  later given your logic of wood colonization or having smaller individual wood pieces or more than one in the agar in the slant? Just a thought for thinking ahead


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#14 esculenta

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Posted 06 March 2014 - 10:40 AM

what about making slight perforations in the stick not so it will break but to allow for multiple stick piece transfers  later given your logic of wood colonization or having smaller individual wood pieces or more than one in the agar in the slant? Just a thought for thinking ahead

 

ideally you don't keep slants for much more than a year, so I've heard.  You don't need to take pieces of wood when you transfer.  It just gives something for the myc to chew on for the long run, and is supposed to keep the wood loving enzymes flowing.



#15 Jeff3

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Posted 06 March 2014 - 09:12 PM

what about making slight perforations in the stick not so it will break but to allow for multiple stick piece transfers  later given your logic of wood colonization or having smaller individual wood pieces or more than one in the agar in the slant? Just a thought for thinking ahead

 
ideally you don't keep slants for much more than a year, so I've heard.  You don't need to take pieces of wood when you transfer.  It just gives something for the myc to chew on for the long run, and is supposed to keep the wood loving enzymes flowing.


Exactly. The wood is not your primary source for a transfer.

@ Myc, thanks but the coffee stick is not my idea. I would give credit to the guy that came up with it but I have no clue who it was. :)
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#16 Seeker2be

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Posted 06 March 2014 - 09:16 PM

what I was thinking was a few sticks if colonized could colonize multiple petri dishes instead of just one so breaking them off at perforated sites would make it easier to do sick to petri  transfer as well as with slant agar to petri but I do understand they are there to get the mycelium used to the wood and give nutrition and not to be used to noc up petri dishes.



#17 Jeff3

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Posted 06 March 2014 - 09:39 PM

It may work Seeker. Not knocking your idea.

It is best to have a schedule for backing up your backups though. Every 18 to 24 months take a sample to a petri, grow it out and re-slant it. If you don't have too many it is an easy project to handle. If your dish fails then go to the wood. If the first sample of wood doesn't grow I am not sure that having multiple pieces to choose from is going to be all that beneficial. More than likely it is dead and you are going to need to start over.

Just my two cents. :)
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#18 Seeker2be

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Posted 07 March 2014 - 01:12 AM

Thanks for brain storming with me.  There are still pertubations of this hobby to be discovered I think and that what keeps it exciting. .  I appreciate your input Jef


Edited by Seeker2be, 07 March 2014 - 01:13 AM.

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#19 stoffel

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Posted 07 March 2014 - 06:58 AM

i like agar teks, very good thanks just learned some things new ^^

 

greets

 

stoffel


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#20 Devu

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Posted 21 September 2014 - 03:17 AM

I know this is old but thanks for sharing this.






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