42 replies to this topic

[Jrotten]

My how fast everything turns around... 

 

I emptied my mini fridge of 9 months of hoarding cultures and took out all the clone plates, leaving 10 plates.  One was a recent duplicate.  Today I took 4 transfers of my first clone, a B+ from the first grow when I started back last year.  It was bacterial and I transferred it to a respectable looking couple of plates.  It was a massive flusher I believe because of the bacteria.  It went to antibiotic agar today to test the hypothesis.  Next was the 4 Arenal Volcano cube clones I was working with last spring 2 transfers each.  The are a few generations domesticated, potent, and have a propensity for nipples so I decided they were a good candidate for my first effort at culture improvement.  3 Semperviva clones got 2 transfers each because I have been monkeyfingering instead of whitethumbing none has fruited, though 2 just got cased, fingers crossed.  Finally a pan clone from the last tray that went green got 2 transfers and one plate was left alone to grow.  I've never had full sized pans yet, just dwarves and pins, but the pins were plentiful so I want that culture.

 

Then I put each of these 9 cultures to about one cup in half pint jars (half full) of WBS.  The plan is to micro grow each of these in one of two fruiting chambers alongside some Gymnopilus experiments.  The idea is to use cultures I'm somewhat familiar with to do micro grows now that I have a setup that will facilitate that and work the bugs out of a system I have yet to put together.  Hopefully they all come back, but I expect some won't recover well from 3-6 months in the fridge.

 

THEN I poured a sleeve of 25 plates with a corn syrup/MEA agar with a pinch of coffee grounds and sawdust.  They are cooling now.  When they are cool and I have the time I am swabbing 3 plates each with:

Pan Cambo var Jambo

Zapotecorum from Australia

Subaeruginosa 

Pan Cyan from Texas supposedly an F1

Mexicana Jalisco

Mexican Chincho Nindo

Tampanesis

Semperviva genetics that are new to me

1 plate with my Semperviva spores

 

And just like that, I emptied my fridge and filled my incubator!

[rockyfungus]

Today I went into cold storage and retrieved all my slant cultures. Started thinning out duplicates.

I have a few new strains to add to my library and need to make space! Shit gets out of hand quickly.

Loving the list, mine is more gourmet/albino cubes for now.

[Jrotten]

Today I went into cold storage and retrieved all my slant cultures. Started thinning out duplicates.

I have a few new strains to add to my library and need to make space! Shit gets out of hand quickly.

Loving the list, mine is more gourmet/albino cubes for now.

I'm not nearly as successful a grower as that list would sound, but I'm hoping to be. I just got streaking them all.  It was a good morning to drink my coffee, eat my oatmeal and just go get into the zen of the process.  The Subs and Zapos are supposed to be wild but I have a hard time believing that because the print quality was stellar and HEAVY.  Now I just need to leave everything alone for a week or so and resist the urge to peak!  Hopefully the little grain jars get to moving quickly!

[DrepsiLocybe]

are you using gentamicin as antibiotic agar? if so whats your ratio? ive tried 0.2mg antibiotic to 500ml lme
have been trying to clean up some cultures and also get some old spores back...
got my e4k to take from a 3 year old dried fruit and it took almost 2 months to show growth :)

[Jrotten]

I am and I used 5mg/1000mL per instructions. What I found is that it’s not a miracle at that dose and the bacteria will actually grow for several days. In my case the very bacterial pins I dissected and transferred to antibiotic agar seemed to be worse off after the first few days, but then the bacteria was contained. And the fungus outgrew it, then I transferred then I transferred again plus an inoculation wedge to grain. Sort of to test whether it would need another transfer. I think more might work better, or even a different antibiotic, but if it actually works I think there might be benefit to having to compete with the bacteria for a few days?

The whole process was:
Harvested pin> normal mea> cut pins and tried transferring the most recovered least bacterial sections to Gentamicin agar> transfer when the mycelium had substantially outrun the bacteria> from that totally clean looking plate two transfers plus a grain inoculation.

Edited by Jrotten, 26 November 2020 - 10:28 AM.

[Jrotten]

In 3 days it looks like all my Tampanesis plates are germinating at a very high rate.  One Zapo plate has a spot of myc and spot of bacteria which i expect means that plate is a goner.  Most everything on grain has recovered very fast and is going.  The pan clone is a little slower to recover and the Semperviva clones are all hit or miss.  The cube clones have all come back extremely aggressive some more so than others.

 

I need to get another sleeve or two of plates ready and realistically it's time to put together subs and get containers figured out for the grains.  I might G2G whatever looks best from last year.  Depending on how the next 3-4 days goes i might G2G everything to a quart that is worth the transfer.  Anything that isn't good on grain or agar at that point had it's chance, but I expect to have a lot of agar transfers I have to make in the coming days.  Hopefully germination and colonization stagger the cultures on their own accord.

[Jrotten]

Grain jars: everything but 2/3 of the Semperviva clones has recovered and most have taken off well.

Old cultures: Arenal Cube Clones are all rididculously healthy, B+ is hit or miss, but I have one good plate.  The Semperviva clones on agar match grain.

New Cultures: Everything but Subs and maybe Jalisco seems to show life.  Throwing out one Zapo plate for contamination I believe was caused by the handling and it's the one I can confirm germination on so that's something to wait and see.  The Chichon Nindo plates just came to life which I'm excited for, and perhaps neatest of all is that the lightest print I had which was a Semperviva print so light I could barely tell where to put my swab has come to life heavily and is only behind the Tampanesis which took off on day 3.

 

All in all for 7 days I feel like I'm off to a good start.  All the grain jars that had really started to do something got a shake around 20-25% today.  I'm not trying to stress things too much, but I feel like with this many cultures and this little grain in each jar, a single shake should help thin anything that just wasn't going to make the cut.

 

I believe I am also going to change up my substrate and just go with Don Shadow's coir and coffee recipe except for the pans and wood lovers.  It just seems easier to not have to make so many batches with this many different things growing.  I could easily just make a batch, put it in quarts, sterilize it and put it aside until needed making it easier to break the work up as things progress.

[Jrotten]

I made a lot of transfers and wrapped a sleeve of plates for future use. Things generally looked to be recovering wonderfully.

[Jrotten]

And the staggering continues.  Today 2 of the AVC cube clones went to an experimental coir/coffee/bran/sand substrate.  Probably 1:4 or 1:5 spawn rate which may be too light.  The idea of supplemented coir is new to me and following the recipe I did I wound up with twice as much as I was supposed to which is a bit worrisome so I'm glad at least the cubes are going in first to test.  3 LC were started with a pan cyan clone, a shot in the dark with a pan cyan TX T1, and the semperviva clone from last season that has been growing on agar and grain since coming out of the fridge.

 

2 other AVC clones, the pan clone, and the semperviva clone are all approaching full colonization on grain and spawning.  And I finally made transfers of Chichon Nindo, Pan Jambo, and Zapo.  The Jalisco and Subs have not germinated at all.  I will likely retry the jalisco.  The Subs I tried 3 days in the fridge to see if they'll germinate.  I may just go with a straight MEA instead of corn dextrose/mea mix for anything that didn't germinate well.

[Jrotten]

Did 9 half pint jars and 7 quarts. 3 of each are for LC test and have manure or compost. Of the straight grains I did
Half pints
2 chichon nindo
2 tampanesis
2 pan cambo Jambo
2 Semperviva from Don Shadow
1 Semperviva 2nd gen from my spores

Quarts
3 different Arenal Volcano cubensis clones
1 Tampanesis

For the mixed jars I am using a Semperviva clone, a pan TX, and a pan clone from Choco that I thought petered out, but may have just taken a moment to recover. I’ve never used LC on grain and I’m satisfying my curiosity about very heavy LC inoculation using dry verm/manure to allow for very high rates. It may be an awful idea, but I’ve wanted to try it for quite sometime. The half pints are proportioned so that they can be used as G2G spawn and the culture will be trained for the final substrate. The quarts are closer to a final fruiting substrate proportion, basically direct inoculation, but using the LC to actually provide all hydration to manure/verm.

I threw out the Jalisco and sub plates that never germinated. The Zapo var Western Australia are moving on agar and I think I’ll be fruiting some cube clones in a micro grow test pretty soon.

This whole Tampanesis plate will be used for a quart of grain. The agar experiment with coffee grounds didn’t go great. Everything looks weird on it, but the Tampanesis was least affected. The coffee ultimately did not wind up distributed equally. Here you can see where their is a concentration of grounds by the straight line the mycelium hesitated at.

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[YoshiTrainer]

Wow, things are hoping for you JR, good luck!

[Jrotten]

Thank you. It helps me to keep record of it all but I don’t want to junk up threads until I have something worth sharing.

[Jrotten]

So always learning... using an agar wedge on grain has been a slow process. Leaves room for things to go wrong. I read someone mention that they shake a jar for about 5 minutes after dropping the wedge. It dawned on me how much sense that makes to create many more inoculation points like when a transfer moves around on a plate.

All my cubes, all my Semperviva, all my Pan Cambo Jambo, all the Tampanesis and even the Chinon Nindo which were the smallest imnoculations are 95% on the little half pint jars in 5 days. That’s half the time of just dropping a wedge, waiting, then shaking later on.

So now I know. Single wedges shaken really well are much less likely to contaminate and there’s no real reason to ware wedges. Now I have to increase my timetable. I think G2G will be needed just to buy some time.

Very happy about this discovery and feel like it should be mentioned more prominently!

Edited by Jrotten, 25 December 2020 - 09:59 PM.

[Jrotten]

The Tampanesis quart is pretty great so far at 6 days. I love the way it’s growing off the agar. The half pint jars were 90-100% or better at 5 days. Today I used each half pint G2G to a half quart except the Pan Cambo Jambos which are going to sub. The Semperviva half pint from Don moved the fastest on grain it seems beating everything by a day or so.

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Edited by Jrotten, 26 December 2020 - 09:43 PM.

[Jrotten]

Well, everything that germinated is moving right along. I expanded each half pint of WBS to a quart because I didn’t have time to make dirt at the time. As it turns out the quarts colonized super fast as well and now I have 11 quarts worth of Semperviva, chichon nindo Mexicans, Tampanesis, and cubes ready to spawn. I’ve never worked on this scale and I wasn’t real sure what to do so I figure 1:1 grain:sub and the sub is just 45:45:10 coir, compost, verm. The compost is old subs, cpoo, straw, coffee, some yard trimmings, etc our of my tumbler. The coir was hydrated with 4 quarts run through my French press after I finished the pot this morning. Now I have to figure out containers. This may all be just too much to deal with at once.

I’m worried about contamination before colonization which I seem to have an issue with my containers harboring random spores, I don’t think my spawn is contaminated. Maybe using trash bag liners? I’m considering trying to grow stones from one of the Tampanesis just in the jar. IDK.

Wish me luck. I have 3 gallons of sub pasteurizing in my oven right now hoping that my perfect field capacity holds.

[Jrotten]

Something a bit unexpected that worked and the first fruits I’ve had mature in a very long time is these Pans. I want to say they are a clone culture because that’s what I had to work with, but for the life of me I can’t say for sure. I had a bag of pans from an LC of clone culture that stalled but didn’t seem to contaminate. I cut a slit in the bag just below the seal about two inches to let the bag breathe in my FC and nothing happened. A sniff test told memoir wasn’t bacterial and I never saw contamination. At the same time I had a cup of **I THINK** the same clone from a wedge seem to stick around 95-97%. Shook it and same result. So I dumped the stalled jar in the stalled bag expecting mold and got 100% colonization in days.

I broke the bag down enough to get it in a small 4 cup tray, cased it, and after 12 hours with a lid on put it out to fruit. The first picture is 3 days in fruiting conditions. The second is 24 hours later. Tomorrow is going to be interesting. That mature stipe is as big as a drinking straw and the cap is roughly 2 inches which I find shocking. These are my first mature, non dwarf, non contaminated pans and after months of Semperviva they seem magically fast!

FC is 24 hour full(ish) spectrum LED light (helps to maintain temperature), 74-77F with my fogger/fan set to run up to 95% and fall to 80%. RH. The FC isn’t really sealed and the humidity controller has a lag so it winds up being a positive pressure fog then falling. I have no idea how often it is cycling because checking on it instantly triggers a cycle so I leave it alone.

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[Jrotten]

My first successful (if spotty) pan harvest. 40g wet. I got two dark prints and two light prints from the same two fruits, took two clones samples from the largest fruit and used the lighter prints to swab two plates. I’m quite happy with this outcome. I drenched the tray and hope to get a fuller second flush.

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[Jrotten]

Zapos and a Mexicana Chicon Nindo phenotype that exploded on a plate I had held back went to grain as well as one overgrown plate of Pan Cambo Jambo. That Jambo is so fast and so aggressive I was a little less worried about using a fully grown plate because I think it can out run just about anything for the small amount of grain I used. Made agar to retry with Subs, Pan Tx, and Gym Purp as well as a new allenii print I just received. It’s been strange getting an idea of wood lovers but I’m excited to try a few on plain MEA.

[Jrotten]

One of the Arenal Volcano cube clones I’ve been working with. It was supposed to be a pretty newly collected variety and that happens to be one of my favorite places on the planet. We went frog hunting (just to find them, not to kill them) on LSD one night out on the edge of the rainforest and I’ve climbed as much of that volcano as the law allowed me to. It’s the first mushroom I’ve cloned and worked with towards developing my own cultivar or whatever you want to call selecting and breeding for consistency.

This umbro/nipple phenotype is one of the things I’m selecting for. As far as cubes go these are pretty potent and aggressive colonizers but take a little longer to mature. They are usually pretty large but this is from about 1”x4”x4” of dried sub so they aren’t very tall which I don’t really mind since my FC is crammed full!

In other news the Semperviva are all taking their time fully colonizing, but the Tampanesis and Chicon Nindo have now grown into and through the casing just a bit so I went ahead and gave the casings their first good mistings. I intentionally applied them a little dry. The mystery pans are starting a very sparse second flush, I think the Jambos are due for pins any second now, the clones have recovered but I’m watching for bacteria, and the zapos are taking their time getting going on grain. Fingers crossed!

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[cujoloki]

3 LC were started with a pan cyan clone

 

 

Very interesting and thanks for being a steward of the volcano variety that is new. Right ON! I have a question though. Did you take a cutting from a specimen and just drop it in the LC or was this to agar then LC? If it was from Material to LC, how was this accomplished? Thanks for the thread. Great read!