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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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My Ongoing WIP/Record
#26685865 - 05/21/20 02:45 PM (3 years, 8 months ago) |
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So for people reading this, it's mostly for my own reference and keeping track of things as I go through this study.
That being said, if you decide to look through as I update things and have comments, questions, input, advice, etc, please feel free to chime in. I might be able to figure out how pictures work someday.
So starting out with this, disclaimer, I have had a serious problem with contamination with my half-assed attempts previous to this. So we're going more in depth this time.
To start out, I've decided to set up agar dishes before anything else. I went with a no-pour method to start out with inside rubber-made containers. I went with a straight agar/potato starch mix with my first go-round. 10g of Agar, 5ml of starch, and 500ml of water. I also added some blue food coloring to this batch.
I then cooked them in my PC at 12psi (the max my digital will allow me to put it) for 45 minutes. Hopefully they will work out.
I just got some glass perti dishes, but I haven't made another batch of agar, I plan on making a different recipe if I can find one I like/get a good recommendation.
I have also started prepping my grain spawn, as my PC can only handle so much at one time. I boiled whole oats for 45 minutes before letting them steam off. I then loaded them 2/3rds of the way full in pint sized jars. The jars themselves have a single small hole in the top center, a little less than 1/4". They are covered by two layers of micropore tape. After I set up my jars, I set them up in my PC for 2 hours at 12psi. I then briefly pulled them to cover the tops in tin foil, as they will be stored for about a week, then cooked them for another 2 hours at the same pressure.
My spores should be arriving by Tuesday. I plan on only using them in the agar to start out with.
If you read all this, thanks for sticking around while I basically talk to myself!
-------------------- I live in the middle of nowhere, where nothing grows.
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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UPDATE:
My spores arrives.
I set up my SAB after a careful cleaning process first with bleach and then with Iso.
I set it up on a counter in a small room in which I've had a HEPA filter running for the better part of three days, though I shut it off while I worked.
I set up two of my agar dishes for now in the SAB with the syringe. A fresh set of gloves, and I carefully dripped some samples into my agar before sealing the lids again.
I then removed the Agar from the SAB and set them up in a small container, though there is enough room for gas exchange.
-------------------- I live in the middle of nowhere, where nothing grows.
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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UPDATE:
Current photos of how the Agar is coming along.


-------------------- I live in the middle of nowhere, where nothing grows.
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Bsfixit


Registered: 10/23/19
Posts: 132
Last seen: 2 months, 18 days
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polaritymind
relaxed attention


Registered: 10/10/16
Posts: 994
Loc: Germany
Last seen: 3 months, 8 days
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Wow youre really making sure no contams this time. With this many precautions its definitely gonna work, good luck And I dont see anything I should have to add, you thought of everything so far I think.
Plates look fine too.
-------------------- "to affirm life is to also affirm death" -Albert hofmann
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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First off, thank you very much for popping in an keeping an eye out folks. It both means a lot, and puts my mind at ease a little with working through this kind of thing.
UPDATE:
So yesterday I did a agar drop into 4 pint sized jars. To make sure things went well, I used about half a dish on each jar. I think I poured my agar too thick however as the myc was only spreading on top. Not sure if that's normal, but I'll research it more later.
Last night I also boiled up some more oats. I filled 4 more pint size jars, but this time I used the polyfill method for blocking the holes. I'll compare later, not sure if it will make a difference however.
This time around I saved some of dark liquid left over from cooking the oats. I know people have used this for LC bases before, however I'm not confident in trying that just yet. Instead I used it in my next Agar mix.
This agar mix was 150ml of 'Oat Broth' 50ml of distilled water, 3.5g of gar, 3.5 grams of potato starch, 1 gram of dextrose, 1 gram of yeast.
It seemed like a lot, but I had everything on hand, so I figured worse come to worse I wouldn't roll with it again. Besides, I'll find out within a couple weeks as it is anyway.
After filling the jars I did another 4 hours total of pressure cooking. I found that the four jars nest perfectly around my flask I use for agar, so it too went through 4 hours of PC.
-------------------- I live in the middle of nowhere, where nothing grows.
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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I know I just posted an update, but I wanted a break between them as they talk enough about different things, and the previous post was busy enough without adding even more.
UPDATE:
The following morning after doing my extra long PC run, I pulled my jars directly into my SAB rather than going to the fridge. After all, I have plenty of plates. Note to self, a single plate goes a long way.
I did another agar drop, once again using around half a plate for each pint jar. The process went a little smoother this time I'd like to think, but as always this is probably my highest risk for contam, so I'm still worried I left things uncovered too long, my blade wasn't sterile enough, etc.
I'll be pouring plates with my different agar recipe soon, then immediately transferring from another plate to the center of me new dishes. I look forward to being able to actually plate my sample at the center of the dish, as my most recent run (in the photos above) was done by syringe and the fluid went rather uncontrollably all over the dish, so I wasn't able to effectively track it's growth.
As such, I am instead just going off of looks. I have one sample that it particularly thick and hearty looking, with strong rope-like webbing in part of it. I plan on sampling that, since I haven't been able to find anything on the forums giving better guidance at this time.
This will probably have another longer break before updating at I wait for my grain to start to colonize, and I wait and see if my new agar dishes provide any success.
Thanks again for those that have taken the time to read through, see you again in 5-12 days.
Cheers!
-------------------- I live in the middle of nowhere, where nothing grows.
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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UPDATE:
Sad update this time. While I am currently experiencing a slow colonization in my first set of jars (micropore tape) my second batch is all contaminated.
I had two main differences with this batch. The first: I used polyfill. I have doubts that this is the reason simply based on how many other people use polyfill to great success, which leaves the other option.
I aired out my grain after boiling it. During my first batch I boiled for an hour, strained, and immediately loaded into jars to PC for 4 hours. This batch, I boiled for an hour, spread out to dry (as I had seen in some TEK that dry grain might work better) then loaded. I then PC'd those jars for 4 hours as well (though there was a slight drop in pressure after 2 hours, so I guess maybe three variables?)
Needless to say, all the jars in the batch have the same tell-tale green mold in them, meaning it was very much something to do with the process. Nothing was different about the agar drop either.
Damn.
Good thing I have more upcoming on rotation here in a day or two. Pictures next time of current growth.
-------------------- I live in the middle of nowhere, where nothing grows.
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Biscuits



Registered: 06/29/17
Posts: 162
Loc: In a society
Last seen: 2 years, 7 months
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That's so weird that you are still getting contams, might have to get more specific.
Can you take photos of the agar plates held up to the light, so we aren't fighting against glare? Would make it a lot easier to see if maybe something is hiding in there. The agar looks clean so far and if it is then it's probably not the SAB's fault.
What temperature are you colonizing the jars at? Is the room clean? Can you be much more specific as to how you're doing the agar drops (where do you store the plates, do you clean them before going to the SAB, sterilizing blade in between jars, etc...). If the agar is clean, given how you're PCing everything to death, I'm guessing the contam is happening in the agar drop or ambiently...
Might be time to invest a few dollars into some SFD filters as well...
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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Re: My Ongoing WIP/Record [Re: Biscuits]
#26825923 - 07/15/20 08:06 PM (3 years, 6 months ago) |
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UPDATE: 07/15/2020
Whew. It's been a while hasn't it?
Okay, from the top. About half my jars ended up alright. The others had this grey-green fuzz spreading at the same rate as the fungus before it overtook the jars. Sadly I tossed them before taking some photos.
When I did the next set of agar drops, two out of my four plates had contams. I'm not convinced I need to rebuild the SAB however, I think something was wrong with my bleaching and iso-ing my small blade I was using to cut samples. When I did a G2G transfer to new jars I used a sterilized set of tweezers and have had zero contams. So there's that I guess?
As far as the shoeboxes go, I have had great success. I started three, all at different rates. The first is fruiting, and the other two are coming along nicely so far. I'll drop some pictures. Sorry for the poor quality, I'm in the middle of packing up my house, so things have been crazy.


-------------------- I live in the middle of nowhere, where nothing grows.
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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UPDATE: 07/21/2020
So, possible problem, but I'm not sure. I think everything has stalled? There doesn't seem to be any further progress. To go into more detail...
I got a solid flush after another day and a half, and I harvested most of those on 17th and 18th. I didn't twist and pull, I used a razor and sliced them at the stem. A few fell over from their weight however and ended up pulling up some substrate. There was already a number of pins, and they popped up even more by the end of the day (still the 18th).
Well, now it's been 3 days and they really haven't shown any progress... come to look at it, the two other monotubs don't seem to be spreading out very well either, pretty much stalled.
Is it possible I used too much substrate to grain for a ratio (1 part grain, 2 parts coir)? Do I need to put them through another 'dunk'? Am I missing something else entirely?
The 1st tub, pins, but stalled.

The 2nd tub, spreading a bit, but pretty much stopped over the past three days.

The 3rd tub, only a few days behind the 2nd. No real progress seems to be being made.
-------------------- I live in the middle of nowhere, where nothing grows.
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polaritymind
relaxed attention


Registered: 10/10/16
Posts: 994
Loc: Germany
Last seen: 3 months, 8 days
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Are they getting light? They need that to fruit. Otherwise it could be moisture or a hidden contam, in either way have some patience and soon you will have clarity.
-------------------- "to affirm life is to also affirm death" -Albert hofmann
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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UPDATE: 08/15/20
Well, I think they have all finished fruiting. Aside from one or two pins that pop up and are not growing, I'm seeing nothing, even after de-dunking them.
These all were small shoeboxes, made with a single pint jar apiece. I dehydrated the yield, and the total dry weight ended up being 55g. So this was around 20g a pint of grain.
I have some quart sized spawn started, and I'll test to see of the yield is in any way based off the amount of grain used. Until next time!
-------------------- I live in the middle of nowhere, where nothing grows.
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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Well, I attempted to use the same methods as before on a bulk spawn, a large monotub style. Contams. Contams everywhere. Greenish, and what I believe was the early stages of cobweb mold considering it was growing in every direction and looked like polyfill.
I have been recommended to look at how I hydrate my grain more closely, so stay tuned, I'll be doing everything again it looks like from stage 1. I'll even be dropping new agar as opposed to g2g since I don't trust the last jar I currently have to be contam free.
Also, I'll need to look into why my jars keep stalling. Hum. Progress soon hopefully.
-------------------- I live in the middle of nowhere, where nothing grows.
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Cegda
Mentat



Registered: 06/23/20
Posts: 183
Last seen: 1 year, 2 months
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Quote:
polaritymind said: Are they getting light? They need that to fruit. Otherwise it could be moisture or a hidden contam, in either way have some patience and soon you will have clarity.
Thats inaccurate. They will grow in the dark just fine. Pinning trigger is mainly evaporation and correct temps. My shoeboxes are basically neglect tek... spawn and wait for a flush
2nd flush can take a couple weeks, I just pop the lid back on and check it every few days if that. Sometimes they push the lid off.. oh well
Dra I think you are probably overthinking things. I have been getting 3-4 flushes consistently with a 1:2 spawn to substrate ratio without adding any additional water. By the 3rd I need the space for a fresh box anyway so I dump it outside somewhere (I've harvested a few that grew outside that way).
45 mins on agar sounds extreme, 20-25 should be plenty. Let your grain dry more than you think it should, then a little more and I bet you have better results.
-------------------- “What senses do we lack that we cannot see or hear another world all around us?” -Orange Catholic Bible, -from Frank Herbert's Dune
Edited by Cegda (10/07/20 06:47 PM)
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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Re: My Ongoing WIP/Record [Re: Cegda]
#26974422 - 10/07/20 08:02 PM (3 years, 3 months ago) |
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Thank you everyone for the ongoing input. This really is a great community.
Quote:
Cegda said:
Dra I think you are probably overthinking things. I have been getting 3-4 flushes consistently with a 1:2 spawn to substrate ratio without adding any additional water. By the 3rd I need the space for a fresh box anyway so I dump it outside somewhere (I've harvested a few that grew outside that way).
45 mins on agar sounds extreme, 20-25 should be plenty. Let your grain dry more than you think it should, then a little more and I bet you have better results.
I more than likely am! Overthinking born from some misfortune for sure. I'll start lower times on the agar, though it'll still be in my small digital PC since I'm one of those heathens that do not own a microwave.
I don't think I'll see any of my dumped runs producing anything anywhere on my property due to my being in the mountainous north, but stranger things have happened.
As it stands I think I will prep a larger run of jars, perhaps 20 or so, to have on hand. They store for some time without the need to refrigerate as I understand it. This will also allow for more growth at any given point in time. The large monotub, while tempting due to the possible flush size, is really not required. I now have room for probably 12 rotating smaller shoeboxes, and this should provide me more 'wiggle-room' with contams until I can get better success rates from practice.
-------------------- I live in the middle of nowhere, where nothing grows.
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Cegda
Mentat



Registered: 06/23/20
Posts: 183
Last seen: 1 year, 2 months
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I have room for monos.. but shoeboxes are easy without mods and all your eggs are spread among baskets..
10 shoeboxes with poorer flushes than I see people getting on here has me with more shrooms than I know what to do with
-------------------- “What senses do we lack that we cannot see or hear another world all around us?” -Orange Catholic Bible, -from Frank Herbert's Dune
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Sir Pentinite
Stranger all the time.

Registered: 05/15/19
Posts: 525
Loc: ation Location Location
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Re: My Ongoing WIP/Record [Re: Cegda]
#26974467 - 10/07/20 08:25 PM (3 years, 3 months ago) |
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Yeah, 10 or 12 shoe boxes will keep a hobbyist busy for sure.
I'm thinking either the cultures aren't clean enough when they go to grain or your technique is letting stuff in. Four hours in a PC should lay waste to everything living in those jars.
-------------------- "I thought to myself 'Boy, I'm sure glad there's nobody here to see this because this is exactly the sort of thing that gets people riled-up and they assume you're dying and that something has to be done. Where if you're alone, you know, you either come through it or you die, but in any case you avoid the fuss.'" - Terrence McKenna
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Dracultivation
Probably Imaginary



Registered: 05/19/20
Posts: 64
Loc: Mountains. High Altitude.
Last seen: 2 years, 11 months
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UPDATE -
Well, I suppose a 50% success rate isn't too horrible when considering that I only created two tubs. In all reality It's more than 50% loss as one tub was a good deal larger, but I'm remaining positive. I also had a first for me, a pretty interesting mutation. On a second or third flush I plan on taking a few prints for posterity.

-------------------- I live in the middle of nowhere, where nothing grows.
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