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InvisiblePastywhyteMDiscord
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Re: Rye Soak Water Liquid Culture! [Re: Violet] * 1
    #21015933 - 12/23/14 08:50 AM (10 years, 27 days ago)

Quote:

Violet said:
"Overly nutritious" isn't a problem with starch nutrition from grains like it is with sweets.  That's why we have absolutely no trouble colonizing on grains such as brown rice but too-sweet agar has problems.
Nothing to fear.  Grainwater can pack a huge nutrition punch without detriment.
The way it works is that mycelium colonizes the material because nutrition is there, but it takes the enzymes to convert the more complex starch into the "sugars" it uses for energy.  I believe a grainwater LC would be more ideal for the reasons that we say it's good to not let cultures consume sugars too long, in that they over time can lose their edge for more complex nutrition so-to-speak.




Which is why I started using grainwater for LC to begin with. Sugar based LC always recovered poorly for me. That and being able to throw a jar in with my grains without worry made it my go to for LC. I am rockin a sugar LC right now tho, probably will test it on boxing day. I hope the SITR doesn't let me down like karo or honey do.

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Re: Rye Soak Water Liquid Culture! [Re: Pastywhyte]
    #21015958 - 12/23/14 08:57 AM (10 years, 27 days ago)

and I was called crazy for using GLC....

:rofl2:

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Re: Rye Soak Water Liquid Culture! [Re: blackdust]
    #21016575 - 12/23/14 11:30 AM (10 years, 27 days ago)

Can't wait for your results Ham,

- Gyo


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Re: Rye Soak Water Liquid Culture! [Re: taGyo]
    #21016774 - 12/23/14 12:06 PM (10 years, 27 days ago)

Quote:

blackdust said:
and I was called crazy for using GLC....

:rofl2:




I never called you crazy for using GLC even tho its sub par and can harbour all manner of contams especially bacillus. No I called you an idiot for many other reasons. Then you call me a nazi for using proper sterile tek and the cycle continues :grin:

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Re: Rye Soak Water Liquid Culture! [Re: Pastywhyte]
    #21016864 - 12/23/14 12:23 PM (10 years, 27 days ago)

If you sterilize right, nothing should harbor contams more than anything else....


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Re: Rye Soak Water Liquid Culture! [Re: Violet]
    #21016927 - 12/23/14 12:38 PM (10 years, 27 days ago)

Endospores can easily germinate after a "standard" pc cycle. It just provides a window. I have dealt with high endospore laden grains and places like Aloha medicinals laughs at the idea that 90 min 15 psi is going to kill everything. I no longer do a 90 min cycle because I consider it more of a minimum. I have a large quantity of grain that has a really high endospore count. Took me a while to figure it out cause I was a believer in the 90 min is good enough doctrine. But now I know what to look for and how to beat it everytime.

FYI aloha sterilizes their spawn twice, the second run is done for 4 hours @ 60psi.

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Re: Rye Soak Water Liquid Culture! [Re: Pastywhyte]
    #21017120 - 12/23/14 01:23 PM (10 years, 27 days ago)

With certain grains I found my early-on 40min PC cycle wasn't enough to ensure no bacterial contams, although it was often fine. Upped it to 45, 50 for full-loaded 250mL brown rice, and that did it.  But it's only the ones in the top of the cooker that had the problems, the bottom didn't, closer to the heat source..  So I'm surprised that 45min for an LC alone in the cooker isn't enough for gwlc.  I mean I only sterilize grainwater agar and grain agar poms for like 25 at 15PSI and don't get internal contams.


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Re: Rye Soak Water Liquid Culture! [Re: Violet]
    #21017196 - 12/23/14 01:36 PM (10 years, 27 days ago)

OK well looks like we are comparing apples and oranges again.  45 min is a fine time for gwlc. I just toss it in with my grain cycle cause it won't hurt if you over cook it. But I still stand by people having endospores in their grain a lot more than they think. The first time I noticed them was in a stone jar that was never shook and only inoculated with a single wedge.  By the time the myc reached the final corner I saw some bacillus just starting. Probably took 6 weeks to germinate but it did eventually. The myc colonized it, I spawned to coir tray with no issues and harvested a lot of stones and a few fruits. It never did mold,  but I thought about it a lot. Later experience gave me even more insight to them sneaky bastards.

I thought you were talking about dusts reference to GLC which is fraught with potential vectors endospores being the big one.  My bad.

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Re: Rye Soak Water Liquid Culture! [Re: Pastywhyte]
    #21017219 - 12/23/14 01:41 PM (10 years, 27 days ago)

Oh and I thought glc was referring, in this thread, to gwlc which is the thread's topic.  Got it.

I used to be a big fan of glc.  After one huge disaster trying to use glc to trim time, I now take the opposite stance, tossing it right in with my dislike for LC (though I would be willing to make gwlc like this for expanding edibles)
LI from poms is now my highest suggestion!


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Edited by Violet (12/23/14 01:46 PM)

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Re: Rye Soak Water Liquid Culture! [Re: blackdust]
    #21017636 - 12/23/14 03:25 PM (10 years, 27 days ago)

Quote:

blackdust said:
I always wonder how an LC preforms when left alone compare to sitting on a stir plate as Statmets talks about.



LC by hand (as I call it) is an adopted means for the home cultivator to experiment with liquid culturing techniques. 
When LC's are left to colonize without the aid of a stir plate and magnetic stir bar they form in cloud-like puffballs of liquid mycelium mass formations.

LC's colonized utilizing magnetic stir plates and stir bars colonize the medium faster.
Also, the colonizing mycelium is able to be kept stirred up.
This creates millions of tiny strands of dikaryotic mycelium without using your hands.
LC's intended for colonization on magnetic plates with stir bars are best inoculated with a "slurry" comprised of sterile water in an Eberbach blender, and known, clean culture on agar from a petri dish. 

The Eberbach blender is filled with the desired ml's of water, then sterilized.
Once the water in the Eberbach is sterilized, inoculate the sterile water with multiple wedges of your chosen agar culture using the "wedge and transfer" method.
Cap the blender, then blend the concoction in three short bursts of 3 seconds a piece. 
Inoculate you sterilized liquid medium broths with the concoction using the "free pour" method in your SAB, or in front of the flowhood.


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Re: Rye Soak Water Liquid Culture! [Re: hamloaf]
    #21019991 - 12/24/14 03:01 AM (10 years, 26 days ago)

Here goes the mediums just before being shook, aspired, and inoculated to their receiving grain mediums.



The mediums are wiped down with an alcohol soaked paper towel, then placed directly downstream of the laminar flowhood.
The parafilm is removed.
The media is grasped firmly with both hands.
The media is shaken vigorously and vertically for 15-30 seconds (I shake for 30).

Because I am working in front of a flowhood, the medium is allowed to be absorbed by the filter disk, spilling out all over the lid and vessel.



The media vessel, and filter material are simply wiped with alcohol, then repositioned directly downstream in the sterile air from the flowhood.

Next, the torch is fired up, and the lid to the liquid media vessel is loosened. 
The recycled syringe's water contents are squirted into a seperate container, then the needle flame sterilized.



Next, the lid of the liquid culture container is removed.
The lid is then positioned directly downstream of the laminar flow of air. 

Close attention is payed to where the freshly shaken mycelium is through out the media.
Aiming for the liquid mycelium, the needle is placed into liquid media vessel and the plunger is pulled.
The lid to the liquid culture is replaced as quickly as possible, and tighlty screwed back down.

The needle is quickly wiped free of any culture that gets stuck to the outside of the needle with an alcohol soaked paper towel.
The needle is then quickly flame sterilized, cooled in front of the flowhood, and the syringe cap is replaced.
A small piece of parafilm (1 inch) is folded in half then placed/wrapped around where the syringe cap meets the body.

Nice thick LC's!



My hands are covered with plastic gloves at all times.
The hands are thoroughly, and steadily soaked in 71% iso alcohol, before, during, and after each step in the aforementioned processes.

While performing aspirations, the cap to the syringe is placed directly downstream of the laminar flow with the open end facing the HEPA.



6 syringes of thick liquid culture were aspired all together.
2 from each medium.
They were each labeled "GT 1, sample 1", ect, and so-forth. 

6 grain mediums were inoculated.
Each grain medium was assigned and designated to it's own sample of the liquid culture. 



Over the next few days the cultures will be monitored visually.
Being inspected for recovery time, vigor of growth, and contamination. 
You will be updated as developments are made.
Thanks for checking this out.

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Re: Rye Soak Water Liquid Culture! [Re: hamloaf]
    #21029181 - 12/26/14 06:19 PM (10 years, 23 days ago)

2 days later the liquid mycelium has past the recovery stage and have begun colonizing the sterile grains.



Over the last few months I have been practicing my LC skills after about a 3 year hiatus from doing LC's.  This particular LC will be the 5th time around the world with LC's in the last 2 months.  All successful except for one bag out of round 4 where a light green mold popped up (probably penicillin)  right along the LC's inoculation pattern.  I don't think it was my LC though, because I have 2 other samples from the same LC that are colonizing right along.  I think the contamination was due to the grains in that bag becoming too dry from excessively running the flowhood that week as one measure to clean and combat a mice infestation everyday for that week.  I won the fight with the mice though.  The mice have not returned to my lab in about 4 days, but a few of my grain bags got dried out a little bit.  Said bags were very slow to colonize at first compared to the bags where the grains were slightly wetter.  The bags with the slightly wetter grains are all fully colonized.



These events/symptoms were noticed 3 days ago.  The only adjustments made to the lab was to not run the flowhood so much, and drop the temperature of the lab 7F to create some condensation in the bags.  Now the bags have recovered and are beginning to colonize at normal speed.



Since I have returned to LC's I have been using 1.5 table spoons each of potato dextrose and light malt extract to a quart of water.  Before that I used to just use LME at the same amount, and I am beginning to think about cutting the dextrose again.  The straight LME cultures always had better recovery once they hit the grains.  I think it has something to do with all the sugar in the dextrose that causes the myce to go diabetic or something once it hits the grains.  Either way, ime, recovery and colonization time are much better when using LC's without the dextrose.

So far I am liking this Rye Soak Liquid culture.  I like the look of the recovered mycelium that this culture has created.  The recovered mycelium has a really extravagant and glossy shade of white to it.  I also like the time it took for this culture to recover and begin to colonize the grains.  I also like the way that this culture already has it's own acronym.  Technically, the water used to create the liquid medium for this experiment was from the end of the boil cycle just before taking the grains off of the stove though. 

The reason I used the water from the end of the boil cycle instead of water from just the straight soak was because I noticed that the water at the end of the boil cycle was much darker suggesting more nutrition in the water, plus there was sediment in the water. 

A few things noted about the culture this media created.  It was slower to colonize than my usual recipe.  In those jars before the shake, it LOOKS like there was a lot of myce in there, but there really wasn't.  Once the cultures were shaken, the yield of mycelium was only about 24 CC's of fresh liquid culture from each jar.  My guess as to what caused the slower rate of colonization is due to the extra nutrition of the media from not being diluted. 

For the next round of LC's, I am going to return to the 1.5 tablespoons of LME only per quart of water.  Cutting out the dextrose all together with the addition of nutritional yeast a 4% (.4th of a gram per quart of water) and play with this rye boil water liquid culture recipe a bit more.  If by somehow this rye boil culturing media parallels or exceeds my expectations, or performs as good or better than an LME LC, I am switching to using grain water at the end of the boil cycle as the main choice for liquid culturing media.  The Next 2 RBLC's experiments are going to be diluted, and one is going to have the addition of nutritional yeast.


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Re: Rye Soak Water Liquid Culture! [Re: hamloaf] * 1
    #21029201 - 12/26/14 06:26 PM (10 years, 23 days ago)

Thanks for the report, glad I'm not the only guy playing with this :super:

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Re: Rye Soak Water Liquid Culture! [Re: Pastywhyte]
    #21029318 - 12/26/14 06:55 PM (10 years, 23 days ago)

They might be doing better as they are used to the substrate

This is from Stamets GGMM talking about agar
Quote:

The pH of the above media formulations, after
sterilizing, generally falls between 5.5-6. 8.
This media can be further fortified with the addition
of 3-5 grams of the end-substrate (in most
cases hardwood sawdust) upon which mushrooms
will be produced. If samples of soil or dung are desired,
they first must be pre-boiled for 1-2 hours
before adding to any of the above formulas. One
potential advantage of the addition of these end substrate
components is a significant reduction in
the "lag period". The lag period is seen when
mushroom mycelium encounters unfamiliar components.
(See Leatham & Griffin, 1984; Raaska
1990). This simple step can greatly accelerate
the mushroom life cycle, decreasing the duration
of colonization prior to fruiting.



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Re: Rye Soak Water Liquid Culture! [Re: blackout]
    #21030581 - 12/27/14 01:51 AM (10 years, 23 days ago)

The mycelium doing so good due to being used to the substrate is what I was thinking too + I have read that chapter out of GGMM a hundred times.  :lol:

There is a correlation here going on between cutting out the sugars in LC, and only using light malt extract, and better performance of the culture that I can't explain.

Bulk subs made from LC's with sugar, ime, haven't done the best at fruiting either.  :shrug:

Knowing how much liquid mycelium to inoculate any substrate with that equals, or would be similar to inoculating a substrate with a high grain ratios would help too I think.

IDK, at this point I just have a good feeling about this Rye/Grain Boil Water as a liquid media, and have stared at the boiling water with the grains in it while considering doing this for sometime now.

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Re: Rye Soak Water Liquid Culture! [Re: hamloaf] * 1
    #21041743 - 12/29/14 10:15 PM (10 years, 20 days ago)

:flyhigh:

Edited by shroominmyroom (07/31/17 11:25 AM)

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Re: Rye Soak Water Liquid Culture! [Re: shroominmyroom]
    #21041756 - 12/29/14 10:18 PM (10 years, 20 days ago)

Why dilute it?  I don't entirely understand.
I don't dilute rgs or wbr water for agar, I just let the starchy sediment settle and pour it off, then use straight.


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Re: Rye Soak Water Liquid Culture! [Re: Violet]
    #21041850 - 12/29/14 10:43 PM (10 years, 20 days ago)

Why such a tall pint? wouldn't that make it pretty hard to suck up the myc?


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Re: Rye Soak Water Liquid Culture! [Re: taGyo]
    #21041859 - 12/29/14 10:45 PM (10 years, 20 days ago)

I dilute it cause there is waaaay more nutes than needed and it makes it easy to see through. But I have used it full strength several times with zero issues.

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Re: Rye Soak Water Liquid Culture! [Re: Pastywhyte]
    #21041877 - 12/29/14 10:49 PM (10 years, 20 days ago)

The see-thru point I understand.  I might would dilute it just enough to make that possible.  Brown rice prep water would be perfect and pack a huge punch!


--------------------
Intentionally or not, here in mushcult we are purveyors of love culture and enlightenment movement. Let's try to act like it!

PODS TEK - Growing Invitro with BRF/verm or Grass Seed containers
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