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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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My first time with agar... Yellow stuff?
#23357043 - 06/18/16 07:20 AM (7 years, 7 months ago) |
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Hi guys so I just started my very first agar experiment three days ago, I was super sterile throughout the whole procedure. I used ms spores on these first three agar plates (jars) but one of them I put inside an incubator as opposed to the others I just left it in SAB after inoculation, and the one in the incubator has yellow stuff all over on top, what's it all about?
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23360470 - 06/19/16 10:48 AM (7 years, 7 months ago) |
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Bump. Somebody.
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23362330 - 06/19/16 11:09 PM (7 years, 7 months ago) |
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Bacteria. Could be a filter failure, you weren't as sterile as you thought, or the bacteria cam from the syringe. Either way definitely keep an eye on the other ones.
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: Mad Season]
#23363388 - 06/20/16 10:11 AM (7 years, 7 months ago) |
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Quote:
Mad Season said: Bacteria. Could be a filter failure, you weren't as sterile as you thought, or the bacteria cam from the syringe. Either way definitely keep an eye on the other ones.
Damn, they all three got the same yellow stuff. So there's no way the mycelium could overcome it ? Also I was pretty sterile but what should I do for filter? I just use tin foil atm to cover the 1/4in hole in the lid, I don't have tyvek or any special tape, would polyfill work?
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23364132 - 06/20/16 03:28 PM (7 years, 7 months ago) |
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Yes poly works. I'd just make new jars.
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: Mad Season]
#23369299 - 06/22/16 08:17 AM (7 years, 7 months ago) |
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Quote:
Mad Season said: Yes poly works. I'd just make new jars.
So looking at them today, it seems like mycelium is starting to form, could it just be yellow maybe from the honey?
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23369306 - 06/22/16 08:20 AM (7 years, 7 months ago) |
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-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23369321 - 06/22/16 08:28 AM (7 years, 7 months ago) |
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Okay this is the best looking one, the mycelium seems to be going strong and colonizing over the bacteria looking stuff. I just have like only 1cc of these spores left I want to get some good mycelium going so I can keep this strain alive..
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23369330 - 06/22/16 08:30 AM (7 years, 7 months ago) |
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-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23369408 - 06/22/16 09:15 AM (7 years, 7 months ago) |
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Quote:
ShroomzGod said:
Quote:
Mad Season said: Yes poly works. I'd just make new jars.
So looking at them today, it seems like mycelium is starting to form, could it just be yellow maybe from the honey?
Definitely not. Definitely is bacteria.
Quote:
ShroomzGod said: Okay this is the best looking one, the mycelium seems to be going strong and colonizing over the bacteria looking stuff. I just have like only 1cc of these spores left I want to get some good mycelium going so I can keep this strain alive..
Yes it is possible to save that strain. Just going to be much more difficult. You will have to transfer multiple times until it is clean and organized. Here's a thread I made to show why.
https://www.shroomery.org/forums/showflat.php/Number/22020260
Please read that thread thoroughly. It's best if you transfer a piece asap as well. Also you should check out my improve your sterile techniques thread.
https://www.shroomery.org/forums/showflat.php/Number/21889950
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: Mad Season]
#23374123 - 06/23/16 03:18 PM (7 years, 7 months ago) |
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Okay so here's how it's looking today, plenty of mycelium. Now if I transfer it to another agar plate how exactly will that help clean it, wouldn't I just be transferring the bacteria with it?
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23374130 - 06/23/16 03:19 PM (7 years, 7 months ago) |
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-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23374158 - 06/23/16 03:28 PM (7 years, 7 months ago) |
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Like I said you should have transferred it yesterday.
Anyways yes you will transfer both bacteria and mycelium, just go for the strongest area of myc. Probably for quite a few transfers the bacteria will be riding along. What you want though is the mycelium to outgrow the bacteria post transfer, and transfer a clean wedge before the bacteria catches up. This may take 5+ transfers, and it is fun IMO, but definitely much harder than normal. That's why I recommended you to start from scratch. IMO you can clean up any culture with the proper agar skills.
There's also the agar sandwich tek where you pour hot agar over the plate and shallow transfer the first bit of myc that pokes through the middle. But I'd do that when you have proper lids with poly filters or something. If bacteria grows on the surface, youll be in the same rut.
Edited by Mad Season (06/23/16 04:31 PM)
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ShroomzGod
MysticMusicalMan

Registered: 02/06/16
Posts: 141
Last seen: 6 years, 9 months
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Re: My first time with agar... Yellow stuff? [Re: Mad Season]
#23376550 - 06/24/16 08:10 AM (7 years, 7 months ago) |
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Quote:
Mad Season said: Like I said you should have transferred it yesterday.
Anyways yes you will transfer both bacteria and mycelium, just go for the strongest area of myc. Probably for quite a few transfers the bacteria will be riding along. What you want though is the mycelium to outgrow the bacteria post transfer, and transfer a clean wedge before the bacteria catches up. This may take 5+ transfers, and it is fun IMO, but definitely much harder than normal. That's why I recommended you to start from scratch. IMO you can clean up any culture with the proper agar skills.
There's also the agar sandwich tek where you pour hot agar over the plate and shallow transfer the first bit of myc that pokes through the middle. But I'd do that when you have proper lids with poly filters or something. If bacteria grows on the surface, youll be in the same rut.
Okay my man first off, I'd like to say thank you greatly for all your help so far, you seem really experienced with agar. So I went ahead and knocked up my other three agar jars that I had chilling in the fridge, I put polyfill in these ones, and I used a different innoculent for each jar, one is with LC from the previous strain I been working with, another one I used the last of the spores I had left from new strain I really wanna get going, and finally I did a transfer like you suggested. What I did is I put water in a syringe up to about 8cc mark, I wrapped it in foil and Pced, then under my SAB with the most sterile measures I purged the water into the colonized contamed agar jar, and tried my best to get mycelium into the syringe and made my transfer to the other agar jar. I hope this works, I'm not so sure I was even able to get any mycelium into the syringe, what's a good method for sucking the mycelium off agar wedges?
-------------------- "And it all appeared very real to many of us, but those who were wise enough to look just a little bit closer could easily see it was all just an illusion" -Looking Out at the World Looking In
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Re: My first time with agar... Yellow stuff? [Re: ShroomzGod]
#23376751 - 06/24/16 09:22 AM (7 years, 7 months ago) |
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Quote:
ShroomzGod said: Okay my man first off, I'd like to say thank you greatly for all your help so far, you seem really experienced with agar.
Ty 
Quote:
I went ahead and knocked up my other three agar jars that I had chilling in the fridge, I put polyfill in these ones, and I used a different innoculent for each jar, one is with LC from the previous strain I been working with, another one I used the last of the spores I had left from new strain I really wanna get going, and finally I did a transfer like you suggested.
Nice! 
Quote:
What I did is I put water in a syringe up to about 8cc mark, I wrapped it in foil and Pced, then under my SAB with the most sterile measures I purged the water into the colonized contamed agar jar, and tried my best to get mycelium into the syringe and made my transfer to the other agar jar. I hope this works, I'm not so sure I was even able to get any mycelium into the syringe, what's a good method for sucking the mycelium off agar wedges?
I can honestly say I've never done that lol. But yeah it should have worked. Just not what you want when you are trying to cleanup a culture because you aren't controlling what gets transferred. What I've done is just cut out a wedge that I'd say looks clean with a scalpel and transferred it to a new plate using said scalpel/x-acto with #11 blade. As seen here..
Quote:
Flame scalpel

While flaming the scalpel, you want to simultaneously pop and slide the lid off like this. I find sliding it really helps in keeping the air still:

Cut a wedge in the agar. Make sure not to let your gloved hand hover over the media!

Stab at it at a 45 degree angle and lift it straight out. Again with your hand not over any open media.

pop and slide the lid off the receiving plate you wish to transfer to using the same method as above, while simultaneously taking your transfer. It should only require 1 hand anyway to take the lid off.
Don't put the receiving lid down. Hold it. This next part shouldn't take longer than 5 Seconds and only requires 1 hand
Place it onto the receiving plate and cut into the agar of the receiving plate. Slide/cut a line into the agar. The wedge should stick to the receiving plate and come off. This way you aren't jostling the plate trying to knock the wedge in, AND NOT HAVING A GLOVED HAND GO OVER THE PLATE have I said not to let anything go over open media enough times yet? Lol..

Didn't get that? Here's closer pics!

Put the lid on the exact opposite of how you took it off. Slide it over and pop it back on (slide and pop)

The entire write-up is here: https://www.shroomery.org/forums/showflat.php/Number/21889950
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Mad Season
hookers and blackjack



Registered: 09/16/12
Posts: 12,666
Loc: Canada
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Re: My first time with agar... Yellow stuff? [Re: Mad Season]
#23377657 - 06/24/16 01:58 PM (7 years, 7 months ago) |
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