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OfflineMaJiK_420
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: ghiajake]
    #19555116 - 02/11/14 11:44 PM (9 years, 11 months ago)

I tried and had one success and one failure with LC before

Kinda curios to try out this way, I have enough LME to last years with Agar, would that work for LC?

And I think I remember reading about no tilt jars before, but I can't remember why make it no tilt?


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Offlineghiajake
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: MaJiK_420]
    #19555140 - 02/11/14 11:49 PM (9 years, 11 months ago)

Yes, lme is perfect. Use .6g of it for every 100mL of water, like I said in the OP. The no-tilt lids are much easier to extract from, you can use both hands on the syringe instead of holding the jar and one working the syringe. They aren't completely necessary, but I highly suggest them.


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Offlinex7x_x7x
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Re: Liquid Culture De-Mystified: Ghia Style [Re: ghiajake]
    #19556839 - 02/12/14 11:22 AM (9 years, 11 months ago)

Quote:


    Now, enough bragging, let's get down to business making some LC! Keep in mind, LC is meant for live culture transfers only!!! Trying to germinate spores in an LC solution will end in failure 9.9 times out of 10.




i germinated spores in LC many times and always work ok


--------------------
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koh samui and oak ridge are my favourite strains


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Offlinex7x_x7x
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Re: Liquid Culture De-Mystified: Ghia Style [Re: x7x_x7x]
    #19556845 - 02/12/14 11:23 AM (9 years, 11 months ago)



--------------------
cultivando en la miseria

SuctoSpore® Pictorial Tek


x7x_x7x@shroomery.org

carl_jung_in_lsd@yahoo.com

koh samui and oak ridge are my favourite strains


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OfflinecronicrFacebook
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: x7x_x7x]
    #19556846 - 02/12/14 11:24 AM (9 years, 11 months ago)

insulated lunch boxes workk great:cool: i take mine out and leave it for the day before opening it


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OfflineiFung
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: cronicr]
    #19556858 - 02/12/14 11:27 AM (9 years, 11 months ago)

I thought they sold spores as opposed to LC because spores are legal to sell while LC isn't? (at least among active strains)


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Offlineghiajake
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: iFung]
    #19556909 - 02/12/14 11:46 AM (9 years, 11 months ago)

Quote:

i germinated spores in LC many times and always work ok




Awesome, I'll read the link in a minute. Doesn't concern me much since I don't grow actives, but I always love learning new shit.

Quote:

Insulated lunch boxes workk great:cool:




Thanks, I'll try that. Have to find one that'll fit right in there. Not too concerned, since I'm gonna probably have to get a bigger fridge anyways once it warms up enough I can't just use the shelf in my basement anymore.

Quote:

I thought they sold spores as opposed to LC because spores are legal to sell while LC isn't?(at least among active strains)




That is correct, but the OP was written to cover mostly edible/medicinal species. It works just as well with actives too.


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OfflineSaint
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: ghiajake]
    #19566951 - 02/14/14 03:58 PM (9 years, 11 months ago)

i did find more info on my GLC thanks  for looking Grain Liquid Culture (GLC) Tek


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Offlineghiajake
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: Saint]
    #19566971 - 02/14/14 04:04 PM (9 years, 11 months ago)

I think I read that one too.


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Offlinekotter
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: ghiajake]
    #19579309 - 02/17/14 11:13 AM (9 years, 11 months ago)

Thanks for posting that write up about the value of liquid culture.
One huge plus for LC in my world is the speed of preparation if I discover I want to make a clone immediately but don't have any appropriate media already made up.
I can find a nice mushroom in the wild and make up some jars of liquid, have them pc'd, back to room temperature and have bits of fungal tissue in them within no more than a couple of hours.  I've been using this for doing initial isolations for some months now and only going onto agar after I've got some clean looking growth going. Typically I move new growth onto agar at the same time I transfer the rest of the blobs onto some grain and a bit of it into a jar with some fresh LC.  I try to keep a few extra on hand and they also seem to store better than agar. I've left a couple jars sitting around since October just to see how long they will last and neither has contaminated with anything yet.
LC has proven itself to be a valuable approach for me this past year. Or at least, whatever the reason, my success rates for obtaining clean cultures from new clones has gone way up.


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Invisibleblindingleaf
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: kotter]
    #19579340 - 02/17/14 11:21 AM (9 years, 11 months ago)

:whathesaid:

nice write up!

i would add into the OP the option of putting an agar wedge into an LC to start it.  works wonders as well.  thats the only way i do it.  the first time i used grains to noc up an lc, then when i started with agar, i moved to that. 

jake, was wondering why u do not like honey or karo?  i use honey, but am getting some LME in the mail soon.  will substitute that for honey next time, but still curious.


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InvisiblePastywhyteMDiscord
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: blindingleaf]
    #19579373 - 02/17/14 11:31 AM (9 years, 11 months ago)

I don't like LC for cubes at all but, I am a huge fan of it for p galindoi :thumbup: LC  is an excellent choice for slow colonizing species. Just need to do it right.  A large capacity syringe is a must IMO as you chance for contam increases every time you stick the SHIP. Larger syringe, less sticking. Little things like that are overlooked all to often with LC which IMO why it turns into a disaster.

My LC recipe is so simple. Take some grain soak water, run it through a coffee filter, dilute and use. Works fantastic IMO.


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Offlineghiajake
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: Pastywhyte]
    #19579522 - 02/17/14 12:11 PM (9 years, 11 months ago)

Quote:

blindingleaf said:
i would add into the OP the option of putting an agar wedge into an LC to start it.


 

Step 10, second sentence...
Quote:

If you have no problems with agar, then just noc up your LC with it if you like, but I think the following ways are less prone to contamination.




I just haven't had luck with using honey/karo. If it works for you keep using it, but I like the ME.

Quote:

Pastywhyte said:
I Take some grain soak water, run it through a coffee filter, dilute and use. Works fantastic IMO.




I have done it like that in the past. When I was growing cubes I used the soak water from the straight grass seed I was using (mostly for agar, but made a couple LC). I had to severely dilute it to make it light enough to see through. Unfortunately, I hadn't built my flow hood yet and my jars got contamed. (I suck at agar in an SAB)


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Offlineghiajake
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: ghiajake]
    #19583623 - 02/18/14 01:27 AM (9 years, 11 months ago)

Did up 6 more jars of LC tonight. I'll post pics once I see if there's any good growth.


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OfflineRandomFX
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: ghiajake]
    #19593252 - 02/20/14 03:25 AM (9 years, 11 months ago)

way cool.


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OfflineRandomFX
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: RandomFX]
    #19593273 - 02/20/14 03:33 AM (9 years, 11 months ago)

Quote:

Kotter said:I can find a nice mushroom in the wild and make up some jars of liquid, have them pc'd, back to room temperature and have bits of fungal tissue in them within no more than a couple of hours.  I've been using this for doing initial isolations for some months now and only going onto agar after I've got some clean looking growth going.




you used Liquid Culture to isolate? how did you do that? Or do you mean just to initially hopefully weed out like debris or obvious mycelium growth to 'other things', then move to agar to actually isolate?


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Offlinekotter
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: RandomFX]
    #19596426 - 02/20/14 07:20 PM (9 years, 11 months ago)

Yes. I've found it really helpful for getting nice clones going of mostly wild mushrooms. I started doing this in an emergency pinch when finding a really nice hericium when I was leaving town within several hours. In that case I opened the PC while still it was still hot in front of the flow hood and let the airflow cool down the jar.


I'm essentially using liquid media for the initial isolation. If that growth does not come out clean I pitch it. I've learned the hard way that contaminants can lurk inside some of the tissues on hericium so don't trust them to be clean until after I've done two transfers with the last one being agar. I do that even if using agar for the initial isolation although lately I've been using part of the mycelium grown in liquid to inoculate some grain at the same time I'm inoculating agar. There has been very little incidence of contaminations so far.

I'm using a weak sugar solution (examples: 12 gm LME in a liter of distilled water or 4T Munton's malt extract in a liter or 19 grams light Agave nectar in a liter), cooking it 20 minutes in a half pint or pint mason jar with a filter disk lid, opening up a mushroom body in front of the hood, then teasing out a tiny piece that has never experienced the outer world until I tear it open. I usually make some short incisions around the outer portions of the target zone and then tease out bits of tissue with the tip of the scalpel or with a needle forceps. All of that is pretty normal stuff except maybe for my use of liquid instead of agar to start. I typically use a scalpel or a pair of needle forceps to fish out the wads of goo for transfers. In between transfers I drop the entire tool into a jar of 95% EtOH and then flame it until its dull cherry.


Hericium americanum in liquid malt extract and water




Local Hericium corolloides in agave nectar and water




Local Hericium erinaceus in spray-dried light malt extract and water




Local Pleurotus in agave nectar and water. The core was seriously dense and almost solid. It did not want to turn loose of the glass!




Local Trametes in agave nectar and water




A problem I encountered with this early on was mold riding inside of the Hericum tissues. I've come to believe its not always true but is almost always true when the tissue is soggy. I stopped taking samples from any soggy tissue (which means some specimens don't get cloned) and stopped trying to get branch or tip bits to grow and the incidence of mold plummeted to almost nothing.
On a corolloides I feel fortunate if I can glean a single mm x a mm piece of clean tissue but one is all I need.


I'll add that the vast majority of anything I know came from comments on this forum.
I know liquid does have a bad reputation in some people's opinion but I'd like to suggest that some of that might be not understanding some helpful things like never permitting a sample to float and not using compromised tissues.  If any part of it has seen air other than from the flow hood or has touched another part of the mushroom during removal or been compromised by sog it seems likely it is going to require additional work to get it clean. (If liquid is exuded when the sample is being taken its often a waste of time - this is common for erinaceus.) If the tissue is clean and only moist like normal healthy mushroom tissue a tiny bit is all it has been taking. The base of Hericiums has been the easiest place to find that sort of sample but any inner part that is thick enough to be torn open seems to work too.
Obviously I try to get this completed as soon as possible after finding a mushroom. I now try to keep a few jars of liquid made up and on hand at all times just in case something interesting pops up. 


The way I'm doing it is not amplifying like Ghia Jake's sweet write up above but is just another application of the same sort of media.


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Offlineghiajake
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: kotter]
    #19596515 - 02/20/14 07:45 PM (9 years, 11 months ago)

Dude, thanks for posting this! It's awesome. I will be trying this with some wild specimens this year for sure! Have you ever included an autoclavable anti-bacterial agent in LC cloning solution? Do you dip the wild specimen in H2O2 before tearing and taking a sample? This is a great way conserve dishes and reduce waste stream.

This is what I like, other LC believers coming out of the closet and promoting useful applications of this tool. :smile:


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OfflineRandomFX
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: ghiajake]
    #19596834 - 02/20/14 09:08 PM (9 years, 11 months ago)

Quote:

All of that is pretty normal stuff except maybe for my use of liquid instead of agar to start.




ah ok...so I am getting the impression you really use it just to start a culture, not really to isolate. you just get the culture going, hopefully from a clean sample since you are taking the sample from the interior and then you actually isolate on agar later, am I understanding correctly? I have no problem with LC's, as was stated earlier, they are a tool and a great one for mass inoculation since they can easily spread and thin the live culture out more so then a solid on agar, clearly. I would be surprised if professionals did not use them for bulk inoculation of spawn, and in fact most of us do in reality use them when we put agar pucks in deionized water for long term storage. that is clearly really a type of liquid culture, and in many cases the strain can be stored according to papers I've read, I haven't had them that long to attest to it myself for a 20 yr storage, at ambient temperature none the less. I personally say screw slants...but well I do not know. some species may not like that type of LTS. I did read where a huge culture bank did go completely to DDH2O storage in like 1994 or something, and they obviously know what they are doing, so I am fairly safe in the theory of it working pretty well, I think.


Edited by RandomFX (02/20/14 09:13 PM)


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OfflinePussyFart
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Re: Liquid Culture De-Mystified: Ghia Style (moved) [Re: RandomFX]
    #19596857 - 02/20/14 09:13 PM (9 years, 11 months ago)

Quote:

RandomFX said:
I would be surprised if professionals did not use them for bulk inoculation of spawn.



A lot of them don't.

A lot of them G2G, which is safer and quicker in most cases.


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