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Catattack some Agar 3.0


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#901 CatsAndBats

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Posted 18 April 2020 - 01:51 PM

Dumb question, but taking prints from nice clusters helps narrow down genetics? I assumed all MS was random.

Yes, I learned that from mlbjammer (an old mentor and mod that has retired). It makes sense if you think about it.

 

A cluster of fruit has already expressed desirable, healthy genetics. One can assume that its spores are healthier too. It's a long game though, going spore to spore, printing exclusively from clusters over time will give one better than average genetics IMHO.

 

Of course there's a chance that one will end up with a shitty grow due to rando spores mating and crapping out or expressing undesirable phenotypes, but lemme tell you, every print that he ever sent me was bananas!

 

https://mycotopia.ne...-6#entry1270992

 

https://mycotopia.ne...-3#entry1302837

 

 



#902 CatsAndBats

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Posted 18 April 2020 - 01:54 PM

I am definitely speaking about agar work and isolation of mycelium that "I HOPE" have great genetics based on the characteristics and behavior on agar. Isolation of specific genetics like clustering I do with a clone from one of the fruits....yes MS is a wild west shootout. 

 

NOT a dumb question at all....

 

 

He picked up on a hint that I threw in my post. That said, it is a pleasure having you back and you are welcome to field any question on my threads/posts, as we are two clowns from the same circus!

 

 

 

creepy-clown-kid.gif


Edited by CatsAndBats, 18 April 2020 - 01:56 PM.

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#903 Arathu

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Posted 18 April 2020 - 02:04 PM

post-113856-0-69296400-1587236526.jpg

 

You told me a LOOOONG time ago...."post away!" Hahahahaha I'm glad to be back around the good folk......there's a beautiful new class at Topia-U......

 

A

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Edited by Arathu, 18 April 2020 - 02:05 PM.

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#904 Choices

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Posted 18 April 2020 - 04:05 PM

That all just made me feel a little better. Lately I feel like I’ve been brow beaten by the mush gods. I’m just starting out with agar, so far have had ok luck. My first go at spore to agar failed, but grain to agar from same plates is working just fine. So I’m thinking my spores are no good??

#905 CatsAndBats

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Posted 18 April 2020 - 04:34 PM

 

 

 

That all just made me feel a little better. Lately I feel like I’ve been brow beaten by the mush gods. I’m just starting out with agar, so far have had ok luck. My first go at spore to agar failed, but grain to agar from same plates is working just fine. So I’m thinking my spores are no good??

 

There's a lot of reasons that your spores might not have germinated, cross contamination with a disinfectant, age, not enough moisture on the agar surface, etc.

 

What was your technique? How many did you try?

 

Developing well executed agar work technique is something that requires a lot of practice. It's all about muscle memory, analyzing contamination vectors, developing mindful clean work practice, etc. I find that if I'm fucking up and making too many mistakes, it snowballs out of control and I walk away or do some work that isn't as precise. IMHO mood has a lot to do with outcome.

 

I also work in redundancy, meaning if I'm trying to do something important, I'll do 5 plates. Examples include, spore germination, a valued culture, etc.. Other tasks I'll only do a 2-3, any extras that are clean, become inoculant.

 

FYI, here's my agar primer if you don't have time to read 4,068 pages of my other agar threads:

 

https://mycotopia.ne...izes-agar-agar/


Edited by CatsAndBats, 18 April 2020 - 04:34 PM.

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#906 Arathu

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Posted 18 April 2020 - 05:42 PM

Hahahahaha.....4,068 pages .....that's probably accurate too.... :tongue:


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#907 Choices

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Posted 18 April 2020 - 06:14 PM

That all just made me feel a little better. Lately I feel like I’ve been brow beaten by the mush gods. I’m just starting out with agar, so far have had ok luck. My first go at spore to agar failed, but grain to agar from same plates is working just fine. So I’m thinking my spores are no good??

There's a lot of reasons that your spores might not have germinated, cross contamination with a disinfectant, age, not enough moisture on the agar surface, etc.

What was your technique? How many did you try?

Developing well executed agar work technique is something that requires a lot of practice. It's all about muscle memory, analyzing contamination vectors, developing mindful clean work practice, etc. I find that if I'm fucking up and making too many mistakes, it snowballs out of control and I walk away or do some work that isn't as precise. IMHO mood has a lot to do with outcome.

I also work in redundancy, meaning if I'm trying to do something important, I'll do 5 plates. Examples include, spore germination, a valued culture, etc.. Other tasks I'll only do a 2-3, any extras that are clean, become inoculant.

FYI, here's my agar primer if you don't have time to read 4,068 pages of my other agar threads:

https://mycotopia.ne...izes-agar-agar/
Spore print taken to foil, stored in dark cool spot, spore print opened(first time, since print). Loop flame sterilized, cooled on agar, print scraped, loop swiped in “S” pattern, lid placed on and the two married with para film. All work was done in front of FH. Total of 8 plates between 2 strains (4ea).

0eb27bda7a7784cfa0455262c3d2ae40.jpg

Edited by Choices, 18 April 2020 - 06:16 PM.


#908 CatsAndBats

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Posted 18 April 2020 - 08:39 PM

 

 

 

That all just made me feel a little better. Lately I feel like I’ve been brow beaten by the mush gods. I’m just starting out with agar, so far have had ok luck. My first go at spore to agar failed, but grain to agar from same plates is working just fine. So I’m thinking my spores are no good??

There's a lot of reasons that your spores might not have germinated, cross contamination with a disinfectant, age, not enough moisture on the agar surface, etc.

What was your technique? How many did you try?

Developing well executed agar work technique is something that requires a lot of practice. It's all about muscle memory, analyzing contamination vectors, developing mindful clean work practice, etc. I find that if I'm fucking up and making too many mistakes, it snowballs out of control and I walk away or do some work that isn't as precise. IMHO mood has a lot to do with outcome.

I also work in redundancy, meaning if I'm trying to do something important, I'll do 5 plates. Examples include, spore germination, a valued culture, etc.. Other tasks I'll only do a 2-3, any extras that are clean, become inoculant.

FYI, here's my agar primer if you don't have time to read 4,068 pages of my other agar threads:

https://mycotopia.ne...izes-agar-agar/
Spore print taken to foil, stored in dark cool spot, spore print opened(first time, since print). Loop flame sterilized, cooled on agar, print scraped, loop swiped in “S” pattern, lid placed on and the two married with para film. All work was done in front of FH. Total of 8 plates between 2 strains (4ea).


 

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0eb27bda7a7784cfa0455262c3d2ae40.jpg

 

 

Well, based on what I'm seeing and your description, either your laminar flow is way off, or your print was dirty as balls. That's bacterial soup, and I don't see an "S" shape of contamination, it's peppered all over the plate.

 

So I deduce that you either have a really bad print that you scraped over your plate and then swabbed in an "S" shape (the spore scrape should be done behind the clean plate, not in front or over it), or your hood is producing turbulent flow as opposed to laminar flow.

 

What do the other plates look like?

 

 

 

 

FYI to anyone who wants an easy printing tek, here's one that I wrote:

 

https://mycotopia.ne...eptic-printing/

 

I'd change the alcohol to 70%, but now I'm just nitpicking.


Edited by CatsAndBats, 18 April 2020 - 08:45 PM.

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#909 Choices

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Posted 19 April 2020 - 12:41 AM

That all just made me feel a little better. Lately I feel like I’ve been brow beaten by the mush gods. I’m just starting out with agar, so far have had ok luck. My first go at spore to agar failed, but grain to agar from same plates is working just fine. So I’m thinking my spores are no good??

There's a lot of reasons that your spores might not have germinated, cross contamination with a disinfectant, age, not enough moisture on the agar surface, etc.

What was your technique? How many did you try?

Developing well executed agar work technique is something that requires a lot of practice. It's all about muscle memory, analyzing contamination vectors, developing mindful clean work practice, etc. I find that if I'm fucking up and making too many mistakes, it snowballs out of control and I walk away or do some work that isn't as precise. IMHO mood has a lot to do with outcome.

I also work in redundancy, meaning if I'm trying to do something important, I'll do 5 plates. Examples include, spore germination, a valued culture, etc.. Other tasks I'll only do a 2-3, any extras that are clean, become inoculant.

FYI, here's my agar primer if you don't have time to read 4,068 pages of my other agar threads:

https://mycotopia.ne...izes-agar-agar/
Spore print taken to foil, stored in dark cool spot, spore print opened(first time, since print). Loop flame sterilized, cooled on agar, print scraped, loop swiped in “S” pattern, lid placed on and the two married with para film. All work was done in front of FH. Total of 8 plates between 2 strains (4ea).



&&0){for(var>)throw>
0eb27bda7a7784cfa0455262c3d2ae40.jpg


Well, based on what I'm seeing and your description, either your laminar flow is way off, or your print was dirty as balls. That's bacterial soup, and I don't see an "S" shape of contamination, it's peppered all over the plate.
2 of the 4 where S wiped and 2 were wiped and sprinkled. I was testing(trying)the 2. This one is
of the latter.

So I deduce that you either have a really bad print that you scraped over your plate and then swabbed in an "S" shape (the spore scrape should be done behind the clean plate, not in front or over it), or your hood is producing turbulent flow as opposed to laminar flow.

I scrapped on the side. And this particular plate has the loose spores. (I’ve since learned that “less is more”).


What do the other plates look like?

The other jars are a piece of grain and LC solution of APE They are doing just fine.



FYI to anyone who wants an easy printing tek, here's one that I wrote:

https://mycotopia.ne...eptic-printing/

I'd change the alcohol to 70%, but now I'm just nitpicking.


#910 Arathu

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Posted 19 April 2020 - 07:35 AM

Take four fresh plates and open them, one at a time, in front of your flow hood, just like you are going to streak them with a loop but all we are testing is the air stream itself..Then seal them while still in the "sterile" air stream and incubate them for a week. When you start getting clean plates that stay that way you KNOW that you have a good foundation for deliberately growing targeted organisms. After the above can readily be reproduced then we want to start with the various techniques of streaking/striping which help us to determine if our targeted organism is the one that's growing on the agar. Be mindful of all the moves you make and all of the possible vectors of contamination. It's important to do it the same way all of the time.

 

There is definitely a bacterial soup on that plate you're showing here... anyone that has even attempted this work has wound up with plates that look like that. We're ALL learning..

 

This is a part of moving your skills forward. Aseptic technique and work takes disciplined protocol, practice, and attention to details and is well worth your efforts to learn them.

 

As always IMHO......

 

A


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#911 CatsAndBats

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Posted 19 April 2020 - 09:32 AM

Yeah, I still grow bacteria! I also drink while I work, soooo ¯\_(ツ)_/¯


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#912 Choices

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Posted 19 April 2020 - 04:00 PM

Thank you Arathu , I just made up 32 new no pours. I will test that procedure with my hood. . Again thank you for that. You start to loose your vigor when things keep going sideways.

Edited by Choices, 19 April 2020 - 04:03 PM.

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#913 Arathu

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Posted 19 April 2020 - 04:26 PM

Thank you Arathu , I just made up 32 new no pours. I will test that procedure with my hood. . Again thank you for that. You start to loose your vigor when things keep going sideways.

It becomes a challenge to figure out what is causing our "troubles" and how to mitigate such things within our own limits to that. I'm assuming most of us DON'T have budgeting for research.....hahaha I wish......so we learn and we improvise. You'll get the hang of it soon enough. Keep studying and practicing and critiquing/correcting your own work and It will come. YouTube has a wealth of videos that you may find useful. 

 

Here's a cool video that helps in the visualizing process for air flows. It's important to work to understand the mechanics of the tools we use......whatever those may be.

 

[Direct Link]


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#914 CatsAndBats

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Posted 19 April 2020 - 04:35 PM

Agreed, and we all learn differently. I'm a reader, some people like videos, some people dive right into projects (me), some people prepare extensively before starting.

 

 


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#915 Choices

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Posted 19 April 2020 - 04:43 PM

I have a tendency to dive blind, frothy and head first in to my new endeavors. In hind sight I should of spent more time with cakes and my SAB before moving into agar and bulk. But I also learn my best under fire, and with visuals so I’m sure it will all fall into place here sooner than later. Make a new print from this flush and try again. It’s about all I can do. But I really do appreciate the kind words from all the OG’s, we all started from the same place (is wht I tell myself) haha.
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#916 Arathu

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Posted 19 April 2020 - 05:10 PM

Agreed, and we all learn differently. I'm a reader, some people like videos, some people dive right into projects (me), some people prepare extensively before starting.

 

 

To be certain.....I was not implying different either....... :hug: Ultimately it shows up in our fruiting substrates, or somewhere along the way.

 

Doesn't matter how we learn just that we do......sharing it has the potential to make the learning faster..

 

I think the smoke in the hoods in the video is cool...... :tongue:


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#917 CatsAndBats

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Posted 19 April 2020 - 05:14 PM

I have a tendency to dive blind, frothy and head first in to my new endeavors. In hind sight I should of spent more time with cakes and my SAB before moving into agar and bulk. But I also learn my best under fire, and with visuals so I’m sure it will all fall into place here sooner than later. Make a new print from this flush and try again. It’s about all I can do. But I really do appreciate the kind words from all the OG’s, we all started from the same place (is wht I tell myself) haha.

It's never too soon to get into agar!


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#918 Arathu

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Posted 19 April 2020 - 05:36 PM

 

I have a tendency to dive blind, frothy and head first in to my new endeavors. In hind sight I should of spent more time with cakes and my SAB before moving into agar and bulk. But I also learn my best under fire, and with visuals so I’m sure it will all fall into place here sooner than later. Make a new print from this flush and try again. It’s about all I can do. But I really do appreciate the kind words from all the OG’s, we all started from the same place (is wht I tell myself) haha.

It's never too soon to get into agar!

 

Well said, let me know when I can get a shirt...... :biggrin: 


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#919 CatsAndBats

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Posted 19 April 2020 - 10:53 PM

 

 

I have a tendency to dive blind, frothy and head first in to my new endeavors. In hind sight I should of spent more time with cakes and my SAB before moving into agar and bulk. But I also learn my best under fire, and with visuals so I’m sure it will all fall into place here sooner than later. Make a new print from this flush and try again. It’s about all I can do. But I really do appreciate the kind words from all the OG’s, we all started from the same place (is wht I tell myself) haha.

It's never too soon to get into agar!

 

Well said, let me know when I can get a shirt...... :biggrin:

 

 

This is gonna be the shirt:

 

post-147940-0-39803000-1519145866.jpg

 


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#920 Toybox78

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Posted 22 April 2020 - 12:27 AM

When choosing to add color to your agar, will any liquid food coloring work? All I could find was the Kroger brand. b9c9506d2b23154ad711e70b86038fe3.jpg5aba18c34e72228675664e987cfb8442.jpg

Your friendly neighborhood toybox
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