Dapper_Dillinger
Well-Known Member
Cool story bro
Agar bitches
- Thread starter Richard Drysift
- Start date
Richard Drysift
Well-Known Member
Alright alright alright....
So far we got 3 plates that are currently active. I see one has 3 spores growing out another has two and one on another. What’s cool about agar is you can see each individual spore growing out. No question if it is mycelium or not.
Hoping to take a wedge or two from the best of these and transfer to clean plates this week. This time I will make them thicker so if they make it out towards the edge without contams I know they are clean and good to spawn grains with. Then I just do grain 2 grain transfers until I have enough to do a couple tubs. Thinking I’m going to try doing trays in a shotgun chamber as well. Stay tuned y’all good things are coming...
So far we got 3 plates that are currently active. I see one has 3 spores growing out another has two and one on another. What’s cool about agar is you can see each individual spore growing out. No question if it is mycelium or not.
Hoping to take a wedge or two from the best of these and transfer to clean plates this week. This time I will make them thicker so if they make it out towards the edge without contams I know they are clean and good to spawn grains with. Then I just do grain 2 grain transfers until I have enough to do a couple tubs. Thinking I’m going to try doing trays in a shotgun chamber as well. Stay tuned y’all good things are coming...
Budget Buds
Well-Known Member
I Stab and jab for swabs I guess, this was the first time messing with swabs but since from what I hear true albinos are only found in swab form you just gotta deal with it. I think its crazy to see the mycelium climbing up the swab stick in that AA Plate lol. Normally it's agar to grain
Budget Buds
Well-Known Member
I'm aware of this, time and patience will tell if they mingle .
Dapper_Dillinger
Well-Known Member
Absolutely correct from my understanding you need a minimum of 2 spores to fruit
Richard Drysift
Well-Known Member
I try to take wedges only from visibly rhizomorphic samples. So you are saying these are the result of 2 spores joining? Are there such a thing as male and female spores?
I always thought that if it is white it is definitely mycelium; so it is possible to transfer a wedge to grains and it never produces fruit?
Richard Drysift
Well-Known Member
Ugh so here we go.... these are fucked
Probly could try to get a wedge of the rhizomorphic sample in the top pic that isn’t contaminated just yet but the other plates are growing mycelium now. Just one has done nothing so far. There’s still hope...
Probly could try to get a wedge of the rhizomorphic sample in the top pic that isn’t contaminated just yet but the other plates are growing mycelium now. Just one has done nothing so far. There’s still hope...
Budget Buds
Well-Known Member
A single spore has monokaryotic mycelium, when two spores join they produce dikaryotic mycelium. it takes dikaryotic to fruit. a single spore will not produce fruits, That's what @canndo is referring to , I wait for my agar to fully colonize and then take samples from the line between two obvious spores . Never had any issues getting my samples to fruit and never had any spores not mingle on the agar, I do suppose it could happen though, And as far as I know there are no different sexes of spores, it just requires 2 to work .
Richard Drysift
Well-Known Member
Well that’s interesting. Did not know that. Never had mycelium that had been spawned to grains fail to fruit later on but guess that is possible. Not really trying to isolate anything but a clean culture here.
Budget Buds
Well-Known Member
I would take a sample from every container and put them into the same petri dish to fully colonize , then transfer the best looking mycelium from there.I dont buy into the whole rhizomorphic mycelium is the best , I have an A.P.E.R that only produces tomentose mycelium and it does great . I have been trying to isolate a rhizomorphic mycelium sample for some time but have had no luck thus far , It's mainly just for the experience , not cause it's better
Richard Drysift
Well-Known Member
That’s exactly my plan. Made up some freshly sterile plates last night so let’s see what happensI would take a sample from every container and put them into the same petri dish to fully colonize , then transfer the best looking mycelium from there.I dont buy into the whole rhizomorphic mycelium is the best , I have an A.P.E.R that only produces tomentose mycelium and it does great . I have been trying to isolate a rhizomorphic mycelium sample for some time but have had no luck thus far , It's mainly just for the experience , not cause it's better
Richard Drysift
Well-Known Member
All but one of the original plates that were streaked by ms syringe grew mycelium. Tossed out three that were contaminated with what appeared to be blue trichoderma. Cut a wedge of mycelium from each of the three best looking samples and transferred them onto new agar plates. So now we have 5 plates colonizing rapidly.
I use a sterilized exacto knife inside of a SAB to do transfers; flaming the blade each time with a dab torch. I let the agar itself actually cool the blade with a tiny hiss; does not seem to hurt the mycelium. Pics coming soon
I use a sterilized exacto knife inside of a SAB to do transfers; flaming the blade each time with a dab torch. I let the agar itself actually cool the blade with a tiny hiss; does not seem to hurt the mycelium. Pics coming soon
Richard Drysift
Well-Known Member
So far so good....
canndo
Well-Known Member
I would take a sample from every container and put them into the same petri dish to fully colonize , then transfer the best looking mycelium from there.I dont buy into the whole rhizomorphic mycelium is the best , I have an A.P.E.R that only produces tomentose mycelium and it does great . I have been trying to isolate a rhizomorphic mycelium sample for some time but have had no luck thus far , It's mainly just for the experience , not cause it's better
While ryzomorphic mycelium may or may not signify robust features, one can be relatively assured that the ends of those strands in a sector on a plate is a single culture.
The cultures tend to organize themselves and the ropey parts are likely pure where the more tomatose samples may be mixed.
Richard Drysift
Well-Known Member
Not trying to isolate for purity of variety just a clean culture to put onto grains. Is there a chance that the cultures that are growing on agar did not come from a bisporal union? Always thought that mycelium was mycelium; if it’s all white it’s alright.... right?
canndo
Well-Known Member
W
Wrong.
But the odds of your actually getting the results of a single spore are pretty slim unless that is what you are intending.
I have to dilute solutions to hundreds to one in order to maybe have that happen.
And they DO have sexes, in other words, two random spores may not deliver the clamp connections you need for fruit.
Hey, this is esoterica here. Even the slightest drop from a syringe will have hundreds of thousands of spores and it is a numerical impossibility that two mating spores will not be in the mix.
If you are looking to produce a hybrid then as I said, the opposite problem presents itself.
Wrong.
But the odds of your actually getting the results of a single spore are pretty slim unless that is what you are intending.
I have to dilute solutions to hundreds to one in order to maybe have that happen.
And they DO have sexes, in other words, two random spores may not deliver the clamp connections you need for fruit.
Hey, this is esoterica here. Even the slightest drop from a syringe will have hundreds of thousands of spores and it is a numerical impossibility that two mating spores will not be in the mix.
If you are looking to produce a hybrid then as I said, the opposite problem presents itself.
Richard Drysift
Well-Known Member
Well we shall see what happens then. Never had a culture that did not fruit after successfully colonizing grains.
Richard Drysift
Well-Known Member
So all the transfers look good so far and a few have just about reached the edges; time to prepare the grains. It is always a good idea to make a few more plates and transfer any extra cultures to new plate in order to keep them in play just in case things go south later on down the road. This is another reason streaking agar is superior to innoculating directly to substrate. All you need is one clean “wedge” of mycelium to innoculate a jar of grains and then you can transfer the colonized grain to other jars. Each jar of grain can quickly become 10 more.
For choice in grain spawn many opt for rye berries but I find them expensive; others prefer wild bird seed but some brands got a lot of weird stuff in it like nuts. I like to use whole oats which I get from a local feed store; very cheap at $15 for a 50 lb bag. Been using the same bag of grains for years. Kept dry it seems to last forever.
To prepare the grain I begin by rinsing it the kitchen sink in big ass pot with a lid. 5 jars of dry grain will make 8 or 9 jars of spawn when finished; moisture expands their size. Rinse several times until the cold water looks clear removing any dirt or debris; leave about 2-3 inches of waterline above the grain. Put on the lid and soak the grain overnight. Next day put the pot on the stove and bring to a simmer. Simmer 20 mins and strain off the water.
Allow the grain to fully dry. Wet grains contam easily; use the paper towel test. Spread the grains out on a clean towel in a large shallow bowl to let it steam off while slowly mixing with a slotted spoon. This takes awhile so be patient. Leave it to dry fit an hour and mix again. Do not fill the jars with grain until it is dry enough to put onto a paper towel without spotting. You want “field capacity“ internal moisture but the exterior of the grain should be totally dry. Fill the jars 3/4 full.
The grain jars must have a filter disc, polyfil, or micro pore tape covering a 1/4” hole in the lid for gas exchange. I use polyfil because it is a super cheap, reusable, and effective filter. Cover each jar halfway from the lid with foil to prevent water from getting inside the jars. Sterilize the grain jars along with an exacto knife or scalpel wrapped in foil in a PC at max pressure for 90 mins. Allow to cool overnight.
Use a SAB to do your agar transfers flaming the blade each time. I cut each agar plate like a pizza slice and then stab it to pick it up and quickly drop it into the jar. So that’s 4 jars for each plate. I usually pull 2 from each plate to go into jars and then transfer extras to new plates. Jars take about a month or so to fully colonize; breaking them up as they finish.Then they are ready for spawning or grain to grain transfers.
For choice in grain spawn many opt for rye berries but I find them expensive; others prefer wild bird seed but some brands got a lot of weird stuff in it like nuts. I like to use whole oats which I get from a local feed store; very cheap at $15 for a 50 lb bag. Been using the same bag of grains for years. Kept dry it seems to last forever.
To prepare the grain I begin by rinsing it the kitchen sink in big ass pot with a lid. 5 jars of dry grain will make 8 or 9 jars of spawn when finished; moisture expands their size. Rinse several times until the cold water looks clear removing any dirt or debris; leave about 2-3 inches of waterline above the grain. Put on the lid and soak the grain overnight. Next day put the pot on the stove and bring to a simmer. Simmer 20 mins and strain off the water.
Allow the grain to fully dry. Wet grains contam easily; use the paper towel test. Spread the grains out on a clean towel in a large shallow bowl to let it steam off while slowly mixing with a slotted spoon. This takes awhile so be patient. Leave it to dry fit an hour and mix again. Do not fill the jars with grain until it is dry enough to put onto a paper towel without spotting. You want “field capacity“ internal moisture but the exterior of the grain should be totally dry. Fill the jars 3/4 full.
The grain jars must have a filter disc, polyfil, or micro pore tape covering a 1/4” hole in the lid for gas exchange. I use polyfil because it is a super cheap, reusable, and effective filter. Cover each jar halfway from the lid with foil to prevent water from getting inside the jars. Sterilize the grain jars along with an exacto knife or scalpel wrapped in foil in a PC at max pressure for 90 mins. Allow to cool overnight.
Use a SAB to do your agar transfers flaming the blade each time. I cut each agar plate like a pizza slice and then stab it to pick it up and quickly drop it into the jar. So that’s 4 jars for each plate. I usually pull 2 from each plate to go into jars and then transfer extras to new plates. Jars take about a month or so to fully colonize; breaking them up as they finish.Then they are ready for spawning or grain to grain transfers.
canndo
Well-Known Member
A month?
Shouldn't be any more than 10 days.
I appreciate the work you put in here...but.
No need for gas exchange so long as you use half jar.
Be sure you carve out the out side of each plate before you "pizza" it. The region next to the edge of your plate is the most likely to be contaminated. If your sample has grown out contaminate can light on the grown mycelium, be transfered to new substrate and get an even chance of growing after the transfer.
Sencience can be an issue should you do plate to plate transfers for too many iterations. With oysters I find 5 to be about the limit. Seven with clones from general substrates.