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MrMong


Registered: 10/16/13
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Do YOU alter your agar for different stages of cultivation? 1
#26625457 - 04/24/20 07:05 PM (3 years, 9 months ago) |
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Hello there, currently on day 17 waiting for a drop of MS solution of GT to germinate on agar STILL with no life! I've since inoculated more plates with different vials of spores - so not to worry.
The whole 17 day wait has had me trouble shooting my process and got me thinking about agar and how recipes may improve the liklihood of germination - not only that but how one might alter agar in different stages of cultivation e.g. germination/myc growth for transfers/long term storage.
I have a few questions:
1. Does anyone use an agar recipe specifically for germinating spores? I was thinking a slightly elevated water content and nutrient content (to what proportions I'm unsure) could more readily kickstart germination.
2. Does your agar recipe change when you're cleaning myc via transfer? - I've heard lowering nutrient levels can make the myc more rhizomorphic?
3. Does your agar recipe change when making slants for long term storage?
I've done a lot of reading and searching but would appreciate some up to date opinions/experience/words of advice from anyone, thanks for taking time to read this .
17 days is a long time to wait and I genuinely had a dream (may of been a nightmare) regarding agar plates the other night. Here's one of the suspect plates that I stare at for hours on end and play classical music to, to try and persuade the spores into exchanging DNA with each other
 (Erik Satie was playing when this photo was taken)
-------------------- Flick and Nip
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CocaineBuffet
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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26625486 - 04/24/20 07:19 PM (3 years, 9 months ago) |
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No. Also I would streak that drop with an inoculation loop or a sterilized q tip, gives you more areas to find potential growth.
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Sockadin



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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26625492 - 04/24/20 07:22 PM (3 years, 9 months ago) |
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1: you can make softer plates for starting mycelium on agar from a swipe from a spore print.
2: sometime it can take weeks for mycelium to appear. But your drop doesn't seem like it has started evaporating maybe shake the drop around. Also check your gas exchange. 3: yes lower nutrient agar can be used to express rhizomorphic growth for cleaning up a plate.
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MrMong


Registered: 10/16/13
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Re: Do YOU alter your agar for different stages of cultivation? [Re: Sockadin]
#26627446 - 04/25/20 03:15 PM (3 years, 8 months ago) |
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Thanks for the replies I appreciate it.
I wasn't aware the spore solution should evaporate, does that mean spores have to be in contact with the agar to germinate?
What do you mean by check my GE?
Cheers
-------------------- Flick and Nip
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mimir
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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26627462 - 04/25/20 03:20 PM (3 years, 8 months ago) |
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I use a lower concentration of agar when I'm germinating spores, 20 g/L is my usual and I even get down to 10 g/L. It is very fragile at that point but I've found much better success at germinating spores with softer agar.
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Sockadin



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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26628151 - 04/25/20 08:54 PM (3 years, 8 months ago) |
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Gas exchange. I mean that overtime your water in the spore drop should either start to evaporate or blend in with the Mycelium. Are those jars sealed tight? Or is there a gas exchange filter of some kind?
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MrMong


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Re: Do YOU alter your agar for different stages of cultivation? [Re: mimir]
#26629026 - 04/26/20 07:06 AM (3 years, 8 months ago) |
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Sockadin, they're essentially glass pastyplates with GE holes on lids with 3 layers of 3M tape.
Here's a plate from the same batch of agar that has just had Galindoi germinate on it (inoculated 6 days ago), which makes me think it may just be the GT spores that are the problem.
Quote:
mimir said: I use a lower concentration of agar when I'm germinating spores, 20 g/L is my usual and I even get down to 10 g/L. It is very fragile at that point but I've found much better success at germinating spores with softer agar.
I'm going to make 8 more plates today with this concentration of agar (I'll go for 15 g/L) and innoc them all with these suspect GT spores. Do you think I'll be able to use an inoculation loop on this soft agar without breaking it? If not i'll just shake the drop of spore solution around the surface to spread the spores like CocaineBuffet suggested.
Will keep this updated and hopefully will get some viable GT myc soon!. Cheers guys .
-------------------- Flick and Nip
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A.k.a
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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26629038 - 04/26/20 07:13 AM (3 years, 8 months ago) |
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Yep.
I use soft high nute plates for spores. I know people have spores go off after weeks but 95% of mine grow by day 7 or not at all. I also had harder low nute plates I streaked for lagm which was my first time using spores, and got zero germination. I Reswiped the same prints on soft high nute plates and almost all germinated by day 4.
For early transfers I use regular agar low nute so the myc spreads fast. If I’m keeping one for longer term I make it super nutritious to slow it down.
Once I have clean growth I’ll make a low nute ultra soft plate to use as li to knock up jars. I’d rather drop a wedge that dissolves when I shake rather than do all the blender stuff right now.
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LAGM2020     
Edited by A.k.a (04/26/20 07:14 AM)
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bodhisatta 
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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26629069 - 04/26/20 07:38 AM (3 years, 8 months ago) |
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No. I don't even switch it up usually. 90% of my agar has been MEA
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MrMong


Registered: 10/16/13
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Re: Do YOU alter your agar for different stages of cultivation? [Re: bodhisatta]
#26629364 - 04/26/20 09:59 AM (3 years, 8 months ago) |
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A.K.A that was my thinking, it's very interesting to hear your approach. The ultra soft low nute agar for inoculation is such a good idea! Do you follow any specific recipes for each type of agar?
Quote:
bodhisatta said: No. I don't even switch it up usually. 90% of my agar has been MEA
Interesting, I guess it's not essential to alter the recipe for germination - I'm just stuck with these 2 vials of seemingly sleepy GT spores and want to wake them up!!!!!
-------------------- Flick and Nip
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Sockadin



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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong] 1
#26629888 - 04/26/20 01:24 PM (3 years, 8 months ago) |
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To be fair Bod didn't say he didn't change the recipe. Just that all his plates are MAE... Haha
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cronicr



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Re: Do YOU alter your agar for different stages of cultivation? [Re: Sockadin]
#26629932 - 04/26/20 01:37 PM (3 years, 8 months ago) |
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I don't switch anything unless I run out of ingredients , I use brf but right now I'm out so I'm using millet flour.
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  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
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mimir
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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26631719 - 04/27/20 08:52 AM (3 years, 8 months ago) |
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Quote:
MrMong said: I'm going to make 8 more plates today with this concentration of agar (I'll go for 15 g/L) and innoc them all with these suspect GT spores. Do you think I'll be able to use an inoculation loop on this soft agar without breaking it? If not i'll just shake the drop of spore solution around the surface to spread the spores like CocaineBuffet suggested.
About the recipe, I once put a transfer on a piece of cardboard and it still worked. Mushrooms will grow on pretty much anything, I doubt the recipe is that important. About the percentage of agar though, I just put some spores to half strenght agar today and was surprised by the stiffness. I just opened and new pack of agar from a different brand, so I think it depends on the agar you have. Try it out, if you don't get any germination within 7 days make it softer than before.
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MrMong


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Re: Do YOU alter your agar for different stages of cultivation? [Re: mimir]
#26632289 - 04/27/20 02:27 PM (3 years, 8 months ago) |
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You're right mushrooms are pretty damn adaptable. Normally I don't worry about such fine details but more recently I've been trying to document for the sake of replication and consistency - and paying for spores to not germinate really gets the cogs turning.
I'm keen to learn about other peoples approaches and the biology of spore germination, after staring at empty plates for so long lol.
Inoculated another 8 plates with adjusted agar recipe (thanks mimir) (4g LME: 2g agar (10 g/L), 200ml) with 2 drops of the same GT solution. I gently shook/tilted the plates to spread the solution around. If one doesn't show life in the next 7 days I'll have to buy some more spores
Here's the sexy fuckers before they wen't in the PC - I have a good feeling about them.
-------------------- Flick and Nip
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mimir
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Re: Do YOU alter your agar for different stages of cultivation? [Re: MrMong]
#26634042 - 04/28/20 10:21 AM (3 years, 8 months ago) |
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Great! Let us know
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verum subsequentis
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Re: Do YOU alter your agar for different stages of cultivation? [Re: mimir]
#26634141 - 04/28/20 10:58 AM (3 years, 8 months ago) |
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MEA for me. Dunt change a thing. Every one in a while I'll make it a tad sorry for spores but generally don't even bother.
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