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Shr00merN00ber
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18 Petris, Frist Timer some insight needed!
#26411003 - 01/01/20 01:54 PM (4 years, 1 month ago) |
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Hey guys, happy NY to all! So here we go, my first time doing this properly. I've got 18 petris going. I took tissue samples from the best shrooms growing out of a growbox. There were 3 initial samples, I took a tissue sample from a big mushroom growing out of a cluster hereafter called 'STR', and two different tissue samples from the biggest mushroom hereafter called 'CL1' and 'CL2'. All these dishes are T1 after mycelium grew out from the initial samples where I tried to choose the best sectors.
So I have 2 parts to this post.
1) My pressure cooker only fits 2 quart-sized jars... I'm following this tek, is it up to date? https://www.shroomery.org/forums/showflat.php/Number/20048765 What is the preferred option, prep enough grains for 2 jars at a time so there is no waiting after grains are prepped and pressure cooked? Basically, can I load prepped grains in jars and wait a couple of days till the pressure cooker gets through 2 jars at a time?
2) I would like to know your opinion on the plates. I the dishes numbered written on the dish itself. Is the dish G2G worthy? Single grain jar worthy? Further agar transfer needed? Trash worthy?
STR (4 plates)

CL2 (6 plates)

CL 1 (9 plates <I put two photos of #4 because imo could be the best of the lot, maybe good for G2G?>)

Cheers all, happy New Year again!
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A.k.a
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26411011 - 01/01/20 02:03 PM (4 years, 1 month ago) |
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You should probably clear those lids up then repost.
They look decent from what I can tell. Just set something warm on top of the dish for a min then take a picture.
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LAGM2020     
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Kimble
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26411033 - 01/01/20 02:20 PM (4 years, 1 month ago) |
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I personally would do at least another transfer or two as you are looking for healthy uniform appearance.
I'll mention I found filtering the agar mixture before putting it in the pc can reduce the bits of non homogeneous agar that give the plates a spotty appearance. This helps to give a nice clear view of the culture. Also pouring them a wee bit thinner can also help give a nice clear uniform look.
All that said, they look good. No obvious contams I can see. You're on the right track.
I hope that helps. 👍
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Shr00merN00ber
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Re: 18 Petris, Frist Timer some insight needed! [Re: Kimble]
#26411751 - 01/02/20 12:59 AM (4 years, 1 month ago) |
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Thanks for the input. Regarding the grains, what shall I do? My pc only fits 2 jars at a time.. Can I leave jars with prepped grains waiting? I'm thinking to do the following:
1. Start rinse/soak for about 16 hours. 2. Boil 10 mins, strain, dry and load jars. 3. Start pc with 2 jars 90 mins@15psi, the other jars left in sab. 4. Wait another hour? for jars to cool in pc and repeat pc with next 2 jars and so on.
Will I be risking contams if I do say 10 jars? That will be about 10 hours for the last two jars sitting around before being pc.
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Randalf the Grey
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26411785 - 01/02/20 02:04 AM (4 years, 1 month ago) |
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A 10 hour wait won't destroy your grain. Best case would be the seal\wrap jars and keep them in refrigerator until you are ready to of them.
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Shr00merN00ber
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Thanks for the reply. So after drying grains, load them in jars, seal and leave in fridge till their turn in the pc? Or do you mean after pc store in fridge till they get inoculated later on that day?
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Randalf the Grey
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26411794 - 01/02/20 02:27 AM (4 years, 1 month ago) |
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In fridge until PC. After PC, sit them in SAB while they cool if they are still hot you you take them out. Keep foil on top until ready to noc up. Once PC'd, there shouldn't be any organisms left in the grain to cause spoilage\rot. They will likely be fine just sitting on a counter but I always try to play it safe. Putting them in SAB just limits the amount of contams the outside of jar and filter will be exposed to while it cools. This is personal experience\prefernce talking so please don't take my word as gospel.
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Shr00merN00ber
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Awesome, thanks so much. Will be starting tomorrow. Any petris you see that you like for g2g or particularly dislike? Also, what do you think is best practice, 2 jars per petri dish or 1?
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Randalf the Grey
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26411832 - 01/02/20 03:13 AM (4 years, 1 month ago) |
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Depends on jar size. I only use pints or half pints for G2G (easier to work with in SAB). I put 2-5 wedges per jar. I can't see well enough to point out sectors to cut but can see that they have some uneven growth. I would do at least 1 more transfer before starting a master grain jar. Get a full plate that shows even across the whole thing and you can get 5-10 jars out of it. If youre in a hurry, just do one jar from the best looking edge of one plate while you do more transfers to be safe.
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Shr00merN00ber
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Awesome, thanks so much Randalf. To be honest, I'm surprised you guys can't see the petris clearly, there are a few that the moisture beads can be considered an issue in determining growth but I think most are pretty clear no? In any case, thanks again dude!
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Randalf the Grey
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26411841 - 01/02/20 03:27 AM (4 years, 1 month ago) |
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Happy to help. I think it's more of a focal length issue than condensation. Camera seems to have focused on plates\agar instead of myc. Just not enough detail to give accurate answers.
Also, my eyes suck!
Edited by Randalf the Grey (01/02/20 03:27 AM)
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SpunkyMonkey88
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Quote:
Randalf the Grey said: Depends on jar size. I only use pints or half pints for G2G (easier to work with in SAB). I put 2-5 wedges per jar. I can't see well enough to point out sectors to cut but can see that they have some uneven growth. I would do at least 1 more transfer before starting a master grain jar. Get a full plate that shows even across the whole thing and you can get 5-10 jars out of it. If youre in a hurry, just do one jar from the best looking edge of one plate while you do more transfers to be safe.
What do you mean you put 2-5 wedges per jar?
You dont just cut a plate into 1/4ths or halves and drop a section in?
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Randalf the Grey
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Re: 18 Petris, Frist Timer some insight needed! [Re: SpunkyMonkey88]
#26412266 - 01/02/20 10:30 AM (4 years, 1 month ago) |
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I suppose that is an option, but not how I do it. Each piece of culture that you put in a jar is another point of inoculation. More wedges means more points which means faster colonization. if you have one giant Chunk in a jar and you shake it you're not going to touch as many pieces in the grain as its you have five smaller pieces and shake it. there is another method of using very soft agar. Soft enough that when it is shaken, it almost reliquify eyes and coats all the grain. All of these are options. It also depends a bit on the size of your Petri dishes. You will be able to inoculate more grain jars with a 100 mm petri dish than a 60 mm
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Fonzee
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I'm no pro, but I think you want to take much smaller bits for cloning. Your cultures don't seem too uniform, and thus might require a bit of further isolation before going into mass production with them. The growth seems healthy to me.
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Shr00merN00ber
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Re: 18 Petris, Frist Timer some insight needed! [Re: Fonzee]
#26412329 - 01/02/20 11:06 AM (4 years, 1 month ago) |
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Hey guys, thanks for the input again. The agar sample taken from the initial dish with live mushroom tissue was pretty small, about 1/2 cm. The areas in the dish which seem like an empty circle (eg plate 3 from CL1 between 23-2 o clock) will eventually be filled by stronger mycelium?
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verum subsequentis
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Re: 18 Petris, Frist Timer some insight needed! [Re: Shr00merN00ber]
#26412484 - 01/02/20 12:53 PM (4 years, 1 month ago) |
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I don't see any obvious problems but it's impossible to tell if they are clean because the pics aren't good enough.
To clear up a comment above, someone said "they aren't uniform enough". This is a bit of a misnomer. Plates don't have to be uniform to be clean. It's good to go for uniform organized cultures but not always necessary. Cultures with genetics variety can grow all kinds of "not uniform" and still be clean.
You asked if any of the cultures are good enough for G2G. This is also a misnomer. G2G means grain to grain.... I think you mean "are any of these plates good enough for A2G (agar 2 grain)". The answer to that is probably but no guarantees.
You can load jars and leave them sitting at room temp for ten hours without any issues. I wouldn't bother wrapping them or fridgeing them. I routinely make up more grain than my pc can handle and do it in batches. In fact, as soon as i get off the toilet I'm gonna go start straining and bagging grain. It'll be enough to fully load my PC twice. One load goes in and the other just sits on the counter until the pc is free.
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footpath
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As far as uniformity goes, isn't it still a good indication if there is obvious obstruction or restriction of growth? Case to case, of course.. for instance, in this case I would say that the growth on all of those plates is well within a uniform growth - maybe not perfectly circular, but uniform and unimpeded nonetheless.
But if you have an almost perfect circle of growth with a very obvious divot in one portion of the growth - would you not say that would likely indicate contamination?
I always try to imagine plates in time-lapse as best as I can. I feel like it's kind of aspect of learning to see, if you can take any given snapshot and predict the previous growth process. I feel like those plates lacking in that certain uniformity are ones that you can fairly easily draw back to an earlier date and subsequently diagnose. Maybe I'm crazy though.
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verum subsequentis
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Re: 18 Petris, Frist Timer some insight needed! [Re: footpath]
#26412602 - 01/02/20 02:01 PM (4 years, 1 month ago) |
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You're not crazy. That all makes sense.
There are just too many possibilities and variables to say yes or no. In general, it's a good idea for noobs to look for uniformity but in reality it's an absurd over simplification. Sometimes that's a lot easier than diagnosing every particular noob plate ever uploaded with shitloads of condensation and bad focal points. But, alas, I can't bring myself to give the simple answer.
Plates can be wildly "un uniform" and still clean. It all depends on what's going on in the particular plate. Very nice and clear pictures are needed for one to help online. And even then, the cultivator simply has to learn to see what's going on.
But, yes, i do agree with everything you stated.
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A.k.a
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I always thought uniformity was a sign that it was clean, but mostly that enough strains were removed that if you grow it out it’ll be super consistent.
It’s insane how much there is to learn once you get into agar
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LAGM2020     
Edited by A.k.a (01/02/20 02:13 PM)
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Shr00merN00ber
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Re: 18 Petris, Frist Timer some insight needed! [Re: A.k.a]
#26412829 - 01/02/20 03:53 PM (4 years, 1 month ago) |
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Thanks for the clarification and knowledge verum. I'll keep you guys posted and upload some better pics. Going for the first plate to 2 quarts of rye in my SAB to see whats up. I think I'm going to divide the mycelium into 6 parts, 3 per jar. Most likely going for CL2 #4. Just smoked a fat one too, really excited!
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