Amuseman said:

Just keep transferring until you've got a monoculture, looks good so far!

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mushpunx said:

Amuseman said:

Just keep transferring until you've got a monoculture, looks good so far!

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I wouldn't do this. I would just transfer until you have a clean culture to use as inoculant.


If you transfer all the way down to a single monoculture, you will likely end up with a dud.
When doing strain isolation, after the culture starts to show sectors (typically 8-10 transfers in), you want to transfer each sector to a new plate, and then transfer those until they are monocultures. You want to take from many sectors, so you end up with 10-20 isolates from the one culture, which will then be fruited, tested, and compared. The best isolate, if there even is a solid producer, is the one you keep.

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10kVisions said:

mushpunx said:

Amuseman said:

Just keep transferring until you've got a monoculture, looks good so far!

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I wouldn't do this. I would just transfer until you have a clean culture to use as inoculant.


If you transfer all the way down to a single monoculture, you will likely end up with a dud.
When doing strain isolation, after the culture starts to show sectors (typically 8-10 transfers in), you want to transfer each sector to a new plate, and then transfer those until they are monocultures. You want to take from many sectors, so you end up with 10-20 isolates from the one culture, which will then be fruited, tested, and compared. The best isolate, if there even is a solid producer, is the one you keep.

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Thanks for an educated reply.  I've tried using the loop to spread but after 30 days (10 cultures) nothing happened.  they were fresh syringes and great looking spore count inside the syringe so i was confused there.

how long is the normal wait time to see "good growth" on a culture?

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and now this is the following day photos...



every jar did this.  I'm using a SAB and soaking my knifes in alcohol between transfers as well as heating the blade red hot between every transfer.  so new question is...

if i exposed green mold inside the SAB would that contaminate every jar i open from there out?  would i need to re-clean the SAB between transfers?

also how long can agar jars sit before you use them?  i made my transfer jars about 2 days before i used them and put them in my room just to acclimate to the temp etc.  did i wait to long?

i guess i'm confused as well what im supposed to look for as far as "good" growth is concerned.  super white and fluffy seems to not be good unless i went wrong somewhere.  the "ring" i was getting seems to not be good...

after thought is if "the ring" isn't good and even though i soaked and burnt the knives between transfers and heated it red hot, could it have contaminated the white fluffy jars? i did the ring transfers first then did teh solid white ones... trying to narrow this down here.

mushpunx said:

Those are all extremely contaminated. Toss them all and start again. It looks like they were all mold, or myc very infected with mold/bacteria.
I would give your SAB a good wash with dish soap just because mold.

Dont wash your scalpel with ISO between transfers. It won't hurt anything if you flame sterilize afterwards but it's pointless.

You agar can sit out until it dries out or contaminates. If you don't see any contams it's fine to use. 

Go back to spore. If you used spore solution, place a single drop on the plate, and then swipe it in a zig zag pattern across the plate with a flame sterilized loop. It will make identifying clean myc easier.

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mushpunx said:

You're working in a still air box correct?
The less things blowing in the room the better yea. You need to keep that air still, because you can't keep it sterile

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