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OfflineAce1928
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Grain Agar Hybrid
    #18793212 - 09/03/13 06:05 AM (10 years, 5 months ago)

Hey guys
Uncertain if this belongs here but seeing as it is built upon agar technique and taken a bit further I figured it would

Anyways to the point. I have noticed when transferring from agar to grain, regardless of species, that there is a "lag time" before the mycelium will jump off the agar. Now this annoys me just a little bit. I don't want there to be a "lag" that allows any potential contams to get the upper edge. So I thought to myself about making a grain agar plate.

So I did.

Here is what I did and the first signs of a result

Step one was the normal preperation of your WBS. Soaking. Simmering. All that fun stuff. And i was left with a container of WBS


Now I took 2 empty agar plates and loaded them with essentially a single layer of bird seed


After that I poured in just enough agar to fill in the air spaces but not completely submerge the seeds. So they were like rocks poking out of water


I then stacked them up with the rest of the agar plates that I was also making at the time


And then wrapped them into two stacks of foil and Pressure Cooked them for somewhere between 90 and 100 minutes. WHICH would normally be too much for agar but this agar mixture had very minimal amounts of sugar in it and no caramelisation was evident in any of the plates


Once they were done and cooled I left them for a few days to ensure no contams were left in the seeds. And then transferred some agar from a clone that I made a week or so ago.

This is the growth after approximately 26 hours


So I'm thinking so far this is working quite nicely.

Cheers,

Ace


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Offlinephir3
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18793217 - 09/03/13 06:09 AM (10 years, 5 months ago)

cool, good detail and photos


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OfflineRogerRabbitM
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Re: Grain Agar Hybrid [Re: phir3]
    #18793273 - 09/03/13 06:51 AM (10 years, 5 months ago)

That appears to be horribly contaminated with bacteria and mold.

If you want some grains in your agar, simply grind up a bit and mix it in when you prepare the agar.
RR


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: RogerRabbit]
    #18795811 - 09/03/13 06:57 PM (10 years, 5 months ago)

Hey RR

I can assure you it is not contaminated. I know what you are talking about though. The agar on the plate actually went into the grain and then popped them kind of.

And I have crushed grain into agar before. It does increase the speed a little bit but not as much as an already colonized grain.

This is the next stage from a test I did of transferring sterilized grains on top of a fully colonized agar plate. Those transferred grains were colonized fairly quickly by the mycelium.
I then transferred the grains off of the agar and into a jar and they took off at an amazing speed.

I will keep updating this too.

Cheers

Ace


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OfflineNobitte
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18796396 - 09/03/13 08:54 PM (10 years, 5 months ago)

Im with RR, whenever i supplement grain into my agar, i simply mix some powdered seed in, not that its even necessary most of the time, I've found it makes it harder to see sections for isolation and to differentiate unhealthy/contaminated mycellium, that said, its your time and your call.

Powdering it and suspending it in the agar should also make the nutrients more available to the myc by increasing the surface area and reducing the effort the mycelium has to put in to get access to that delicious, nutritious endosperm.

I love your initiative/enthusiasm, but give powdering it a try.


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: Nobitte]
    #18796668 - 09/03/13 09:36 PM (10 years, 5 months ago)

Few things to reply about that
And thanks for the input

1) All isolation and what not is done on standard agar

2) I don't want the mycelium to have too easy access to the grains. The purpose is to get them making the necessary enzymes for proper grain metabolism firing away before they are in the actual jars

3) Effectively this plate is supposed to act something similar to a G2G transfer rather than agar to grain transfer but on a faster scale and also allowing more individual strains in less space (ie 1 agar plate contains enough grain for multiple G2G and contains an individual strain)

4) I am also finding that it increases the speed that the mycelium acclimatises to the grain. Which is amazing for trickier species such as morels

I have the mycelium from a M. esculenta clone on one plate at the moment (not pictured) which is growing significantly slower than the P. cubensis BUT is starting to take to the grain. This is a good thing because last time it took far too long in the grain jar that it was in and I believe that a bacterial contaminant ended up taking over.

The way I see it is that anything that gives mycelium an edge over contaminants is a good thing and this increases the speed of growth after initial transfer significantly.

PS I'm not claiming that this little experiment will be a blaze of glory, I'm just pointing out what I am hoping to achieve and clarifying a couple of things as well.
I greatly appreciate any and all feedback, positive or negative

Cheers

Ace


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Offlinedrake89
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18799737 - 09/04/13 03:31 PM (10 years, 5 months ago)

Ace: ever try liquid inoculation?  I think you may find it gives you the colonization speed you're looking for.  For me- I use 2 plates per 8oz water in a small mouth pint jar.  Oster blender blades fit in a small mouth mason jar, use the regular metal ring, sterilize.  Blend two petris and I inoculate 4 six pound grain bags with this.  Reishi colonizes 100% at three days.  Others have more lag time.


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: drake89]
    #18800000 - 09/04/13 04:44 PM (10 years, 5 months ago)

I've had really bad luck with liquid inoculation. The few times that I have tried something has gone wrong.
In saying that I have not tried what you suggest in your post though.
And I think I will give it a go when I get some more plates fired up.

One question to you about that particular method with the agar being blended into the water.
How do you ensure that you don't over saturate your substrate?
Say for example you were going to put 4CC of your LC into your jars would you effectively use 4CC less water in the prep of the jars or?

Cheers

Ace


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18800290 - 09/04/13 05:51 PM (10 years, 5 months ago)

Here are some more pics as promised. Things are progressing very nicely so far





That was after an additional 12 hours from the original. Bringing it to approximately 38 Hours



This is 38 hours of morel growth. Substantially slower growth than the cubensis.






These ones are approximately 50 hours after transfer. Looking very nice at this stage



The underside of the plate has got some mycelial growth showing through also.





These last 2 are the morels after 50 hours.
They have made the leap certainly onto the grain. But they still grow so slow compared to the cubensis.

Cheers

Ace


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Offlinedrake89
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18802969 - 09/05/13 11:33 AM (10 years, 5 months ago)

i do add a bit less water, but large bags are probably more forgiving than jars.  It's really just a time saver for me on the farm, taking one step out of the process.  Agar wedges to grain jars is certainly a tried and true method, and I do enjoy your thorough pictures.:thumbup:


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: drake89]
    #18805278 - 09/05/13 09:29 PM (10 years, 5 months ago)

I am planning on trying that technique with blending the agar up into a liquid culture as soon as my next plate is ready to go.
And as soon as I have room in my incubator and what not.

I don't mind agar wedges to grain. It does work well. One jar that I have going at the moment was exactly that. The other jar was inoculated with 3 sterile grains that I put on the agar first.
There is a fairly substantial difference in growth and that is what led me to trying this out.

I will be uploading 2 new pictures tonight of the plates. Possibly the last picture of the cubensis plate as it is nearly finished up and I will be transferring it to jars tonight/tomorrow.

I would love to have some big grain bags going on myself drake. Next thing to do for sure :P


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Offlinedrake89
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18807425 - 09/06/13 01:10 PM (10 years, 5 months ago)

well they seem to be doing good so far except my pearl which is showing the mean green :mad:

Even the shiitake is showing signs of life, which is great because all I got last time was yeast.  Clearly, clean cultures are a necessity for this route, if you're not certain like me it's a big gamble.


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: drake89]
    #18811002 - 09/07/13 09:50 AM (10 years, 5 months ago)

Hey guys

I forgot to take a final picture of the plate. I thought I had but apparently not although in the 24 hours since the last lot of pictures not much had changed. The grains in the pic that look uncolonised were in fact heavily colonised but were slightly dry at the top.

I transferred that one grain plate into another 5 or 6 jars I think. And the mycelium has jumped already (within 6 hours) onto the grain. So I think I achieved my goal. And it is growing rapidly even within those 6 hours.

I will not take pictures of the jars until 4 days time. I am promising myself that I will be patient and wait. Even though its so exciting looking at them every 12 hours or so and seeing how they are. As I'm sure all of you will know

But on that note I am done on this thread. I think it is safe to say that the grain/agar mix was a good idea and that the mycelium on the plate is much better acclimated to the grain in the jars.

In 4 days I will put up a new thread of the current jars for anyone interested in following the progress further. I will post a link in this thread so that the thread trail is easy to follow.

Cheers guys :smile:

Ace


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Invisiblefastfred
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Re: Grain Agar Hybrid [Re: Ace1928]
    #18814066 - 09/08/13 05:33 AM (10 years, 5 months ago)

MEA = spray dried grain extract agar.

I can't imagine that having big chunks of grain in the agar really helps you in any logical way. (vs G2G or other methods)  I'd say prepare some LC if you want good speed.

-FF


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OfflineAce1928
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Re: Grain Agar Hybrid [Re: fastfred]
    #18814942 - 09/08/13 12:00 PM (10 years, 5 months ago)

The speed boost is incredible. The grain in the agar plate was spread amongst 5 other jars and they are all at approximately 20%+ colonization (except the one really large jar) in a matter of 24 hours. I have never experienced that kind of speed. And it is an isolate.

An additional benefit is the fact that the plate that had it in there had residual myc on the sides. I filled it back up with some sterile grains and they too are colonizing very quickly.
It's almost like a mini G2G factory in the palm of your hand.
I'm enjoying using it and the speed is phenomenal.

Less risk than LC. Less hassle than using an entire jar for G2G. Faster than just agar by itself. Not complicated to set up or use.

Regardless if anyone else does this or not I am enjoying using it, and will continue to, and would personally recommend people to give it a try.


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