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Offlinelindens
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Spore syringe on agar
    #26077628 - 06/27/19 03:55 PM (4 years, 6 months ago)

Using spore syringes of GT on agar, this is the result after 4 days.
I'm not sure how much you're supposed to squirt onto the agar but i wanted to get one of those black dots in each dish as seen in the first pic:

in the second pic i wasn't so lucky, only the clear liquid of the syringe made it in:

Are the black dots the actual GT spore? Do these look fine or is it too early to tell?


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OfflineTrichFacebook
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Re: Spore syringe on agar [Re: lindens]
    #26077640 - 06/27/19 04:00 PM (4 years, 6 months ago)

Those both look contaminated- or at the very least, bacterial. Keep in mind from spore to agar it typically takes 4-7 days to see mycelial growth.

The black dots are clumps of spores. A shit load of spores stuck to one another. A single spore isn't visible to the human eye.

The more you put on the plate, the higher the chances you are going to have to deal with contamination vectors.

Next time, use one drop, or as little of the solution as possible.
That way, there's less genetics, and contaminants to siphon through when transfer time comes.


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Re: Spore syringe on agar [Re: lindens]
    #26077649 - 06/27/19 04:07 PM (4 years, 6 months ago)

Quote:

lindens said:
Using spore syringes of GT on agar, this is the result after 4 days.
I'm not sure how much you're supposed to squirt onto the agar but i wanted to get one of those black dots in each dish as seen in the first pic:

in the second pic i wasn't so lucky, only the clear liquid of the syringe made it in:

Are the black dots the actual GT spore? Do these look fine or is it too early to tell?



Those plates are nothing but bacteria! :uhoh:

One drop of spore syringe solution is plenty!


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Offlinelindens
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Re: Spore syringe on agar [Re: Trich]
    #26077668 - 06/27/19 04:18 PM (4 years, 6 months ago)

Thanks a lot! Will set up a few more plates with 1 drop of the syringe this time, and shake up that syringe to disperse the spores.

Another question, the agar i am using was made about 2 months ago (haven't had to chance to move forward until now), pressure cooked and kept sealed in a pyrex bottle (then gelled it up again in a microwave), is it possible it could've expired and is a source of the bacteria?


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Re: Spore syringe on agar [Re: lindens]
    #26077701 - 06/27/19 04:29 PM (4 years, 6 months ago)

A trick I do with spore syringes is after I shake em up real good I try to let some of it settle down near the needle hole....probably doesn't do shit, but it seems to work for me :lol:

Flame sterilize then hold the needle down and let the spores settle then...(drip) :thumbup:


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Offlinek5hd2y
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Re: Spore syringe on agar [Re: lindens]
    #26077707 - 06/27/19 04:30 PM (4 years, 6 months ago)

Doubt it but possible. Make more of it and use an empty plate next time to put your wet spores >loop >agar :goodluck:


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Offlinehexkrak
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Re: Spore syringe on agar [Re: lindens]
    #26077713 - 06/27/19 04:34 PM (4 years, 6 months ago)

Quote:

lindens said:
Thanks a lot! Will set up a few more plates with 1 drop of the syringe this time, and shake up that syringe to disperse the spores.

Another question, the agar i am using was made about 2 months ago (haven't had to chance to move forward until now), pressure cooked and kept sealed in a pyrex bottle (then gelled it up again in a microwave), is it possible it could've expired and is a source of the bacteria?



Depends on the seal on your bottle. That and I'm assuming you didn't leave it sealed when you heated it up in the microwave, so there's that too. However, in this scenario, you're seeing tons of surface bacteria, if it was contaminated in the bottle you'd see bacterial growth throughout the dish, and not just on top. This most likely came from the syringe or happened somewhere during the inoculation/storage process.


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OfflineYasuke


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Re: Spore syringe on agar [Re: k5hd2y]
    #26077721 - 06/27/19 04:38 PM (4 years, 6 months ago)

Quote:

k5hd2y said:
Doubt it but possible. Make more of it and use an empty plate next time to put your wet spores >loop >agar :goodluck:



I one up this. I was having a horrible time with syringe to agar, and once I started just squirting a tiny bit of the inoculant on the inoculation loop and scraping it on agar my success rate went up phenomenally. I dont know if it was the less spore solution or what, but I'm gonna stick with it. I'd have thought the extra contamination factor wouldve made it tougher on me, but nope.


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Offlinelindens
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Re: Spore syringe on agar [Re: Yasuke]
    #26077772 - 06/27/19 05:13 PM (4 years, 6 months ago)

oooo i'll try this as well, i have some fresh loops. When you get it red hot, would a minute suffice for it to cool down before putting some solution on the loop?


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Offlinethe_sonic16
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Re: Spore syringe on agar [Re: Yasuke]
    #26077780 - 06/27/19 05:18 PM (4 years, 6 months ago)

I took the Tormato approach (kind of).

I shook up the syringe and let as much of the visible spores settle as possible. I flamed and squirted a little out while still holding it vertical. I then placed individual droplets one at a time in concentric circles, until the plate was covered in separate circles.

I had to store it with the agar on the bottom, which I know is a no no because of condensation. Luckily, I didn't have any condensate, so it wasn't an issue and I was able to let the drops evaporate and ultimately germinate.


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  • Grain Jar Sterilization - Run 2 - 2.5 hrs on PC
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Offlinek5hd2y
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Re: Spore syringe on agar [Re: lindens]
    #26077781 - 06/27/19 05:18 PM (4 years, 6 months ago)

Never counted,  but yeah I'm sure 60 seconds is plenty.


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OfflineYasuke


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Re: Spore syringe on agar [Re: k5hd2y]
    #26077793 - 06/27/19 05:23 PM (4 years, 6 months ago)

Quote:

lindens said:
oooo i'll try this as well, i have some fresh loops. When you get it red hot, would a minute suffice for it to cool down before putting some solution on the loop?



I put 2 drops right after I flame sterilize it. The first drop is definitely enough to cool it down.
You probably wont be able to see anything on the agar when you inoculate it besides maybe some scratch marks, but its there.


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Re: Spore syringe on agar [Re: lindens]
    #26077800 - 06/27/19 05:27 PM (4 years, 6 months ago)

Quote:

lindens said:
oooo i'll try this as well, i have some fresh loops. When you get it red hot, would a minute suffice for it to cool down before putting some solution on the loop?



A loop will pretty much be cold enough to touch seconds after you remove it from heat. No need to wait


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Before you start...Do you have a Pressure Cooker and a Dehydrator? I highly recommend getting both!


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InvisibleJHOVA
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Re: Spore syringe on agar [Re: Tormato]
    #26077810 - 06/27/19 05:35 PM (4 years, 6 months ago)

yea op should put a drop on a loop and then make a z streak on agar. that way there isnt bacterial water spreading across the entire plate.


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