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Offlinemetalcore_dad
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[FRUSTRATED] Are my syringes contaminated ? What is going on ?
    #26504749 - 02/26/20 11:57 AM (3 years, 10 months ago)

Been at this for 2 months now with nothing to show for it. I have done all that I can to ensure no contaminants. I have SAB, lysol solution everywhere, literally wear a fucking STERILE LAB COAT, flame the fuck out of my needle, and have a PC.


I've been running PF TEK for 2 months literally just trying to grow from spores I bought from vendor, have gone thru 20+ jars, all have not shown any growth whatsoever, or have shown trich contamination. I made a previous post about a test to see whether or not it was my sterile procedure that was causing mold by making a blank jar, and inoculating 4 jars with different syringes. Alongside that, I also inoculated agarose plates with the same syringes to see if mold is present undermicroscope.


RESULTS:


After two week interval:
Blank Jar --> no growth and no contaminants
Jar 1 thru 4 --> no growth, no contaminants


After 6 day interval:
Agarose 1 --> trich mold + mycelium
Agarose 2 --> trich mold + mycelium
Agarose 3 --> no growth, brown puddles, no mycelium
Agarose 4 --> mycelium

I did microscopy for the agarose as well.

Below is mycelium from Plate 4, look at the pretty lil baby hyphae




Plates 1-2 infested with mold, smelled like trich






I need help ... the vendor said the syringes are not "guaranteed uncontaminated" but "no one else has ever said anything like that" but isn't doubting me (obviously is doubting me by saying that). I think the syringes are obviously fucking contiminated. Anybody have input?


Edited by metalcore_dad (02/26/20 12:03 PM)


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Offlineshroomgirl645
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: metalcore_dad]
    #26505418 - 02/26/20 07:31 PM (3 years, 10 months ago)

I'm still new myself, but I've had more luck with pre-sterlyized grain bags, like this one:


if you can't get any luck, buy one of those and noc it up. my first time using them I used a whole syringe, and I just noced up 3 with 1 12 ml syringe, I'll update when that gets going


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Offlinesusurrador
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: shroomgirl645]
    #26505786 - 02/27/20 01:41 AM (3 years, 10 months ago)

That sounds frustrating for sure.

Since you already have the PC, and SAB- why not use agar?

The problem with syringes is that the fruits used to print the spores are probably not grown in sterile conditions. This means that the spore prints that are used to make syringes are not inherently sterile either. Therefore, the syringes are not guaranteed to be 100% clean.

Basically, if you shoot spores straight to grain it will always be a bit of a gamble.

Using agar, you can germinate a single drop from your syringe and see very easily if you're working with mycelium and/or contamination.

With agar you can be 100% once you get the hang of it.

I've used THIS with good success.


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"If it doesn't work, you can always hit him with it."



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InvisiblebodhisattaMDiscordReddit
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: susurrador]
    #26505873 - 02/27/20 04:48 AM (3 years, 10 months ago)

You can't have a sterile lab coat. Unless its in a bag inside a steriliser. Open the bag its no longer sterile.
And copious amounts of lysol is 100% useless.
Take pics of the cakes.
If you got growth on agar but not the cake you make the cakes wrong


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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: susurrador]
    #26506060 - 02/27/20 07:58 AM (3 years, 10 months ago)

I inoculated agar and 100% of them were contaminated. I think there is an issue with the syringes. Thank you for the response :-) Will pursue the agar, perhaps make a liquid inoculant with new spores I get. The entire batch I ordered was from the same spore print so T think it's all contam.


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UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


Edited by metalcore_dad (02/27/20 08:00 AM)


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InvisiblebodhisattaMDiscordReddit
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: metalcore_dad]
    #26506063 - 02/27/20 08:00 AM (3 years, 10 months ago)

I wouldn't say that until you make plates and pretend to inoculate them with an empty syringe and they stay clean. Otherwise you're the culprit not the syringe


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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: bodhisatta]
    #26506071 - 02/27/20 08:06 AM (3 years, 10 months ago)

I've never seen mycelium a single time and have made 20 + jars using PF TEK with PC and SAB.

Here's the most recent jars, 5 days after inoculation. No growth at all.



Also the agar was all 100% contaminated. What now?


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UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: bodhisatta]
    #26506072 - 02/27/20 08:09 AM (3 years, 10 months ago)

I already did that with my jars because I ran a blank jar with no spores, and it didn't get contaminated. I can do that same with agar, where I add spores and then have some blanks as neg con.


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UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


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InvisiblebodhisattaMDiscordReddit
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: metalcore_dad]
    #26506089 - 02/27/20 08:20 AM (3 years, 10 months ago)

Your pf jars should have green mold if the syringes are bad


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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: bodhisatta]
    #26506234 - 02/27/20 10:00 AM (3 years, 10 months ago)

Generally after 1 week of nothing, early powdery mold starts to grow, and then that mold starts to turn green in the center of the white powdery mold. It's trich man. Again, I am as safe as possible, I'll noc some agar and then do some blank agar today but again, I really think it's the syringes.


--------------------
UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


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Offlinesusurrador
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: metalcore_dad]
    #26506318 - 02/27/20 11:03 AM (3 years, 10 months ago)

Sorry OP, I was sleepy and didn't read that you already made some plates.


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"If it doesn't work, you can always hit him with it."



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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: susurrador]
    #26506346 - 02/27/20 11:17 AM (3 years, 10 months ago)

No worries cowboy, I'm just happy my post is getting some attention.

Bodhi suggesting that it still may be my sterile technique but I have a very hard time believing it's me considering it's been 2 months, 20+ jars, and I've never seen mycelium in my jars but saw mycelium in agar, and then 3 days later all of them became overran by mold. It is frustrating.


--------------------
UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


Edited by metalcore_dad (02/27/20 11:18 AM)


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Offlinesusurrador
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: metalcore_dad]
    #26506366 - 02/27/20 11:35 AM (3 years, 10 months ago)

Last winter I spent a lot of time transferring a mold colony around.

Was good practice I guess. :shrug:

When you germinated spores on agar, did you make a streak plate or just squirt the dish?

If you streak the plates using just 1 drop, you're spreading out that drop of spore solution over a bigger surface area and also spreading out whatever is in that drop. Hopefully if you spread the drop out enough, you'll get some mycelium to grow apart from anything else in the syringe and then you can cut a tiny sliver of the newest growth that looks good and move it to a new plate.


--------------------
"If it doesn't work, you can always hit him with it."



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Offlinefeldman114
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: metalcore_dad]
    #26506375 - 02/27/20 11:43 AM (3 years, 10 months ago)

Quote:

metalcore_dad said:
No worries cowboy, I'm just happy my post is getting some attention.

Bodhi suggesting that it still may be my sterile technique but I have a very hard time believing it's me considering it's been 2 months, 20+ jars, and I've never seen mycelium in my jars but saw mycelium in agar, and then 3 days later all of them became overran by mold. It is frustrating.




Are you sure you saw myc?

Sometimes, bunk syringe spores donโ€™t germinate at all.


Also, how did you knock your plates up? Dropped solution on them or used a q-tip?
Only MS plates that ever tammed on me were ones I put an actual drop of solution on. Try the swab method if you havenโ€™t already.


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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: susurrador]
    #26506398 - 02/27/20 12:09 PM (3 years, 10 months ago)

One or two drops, no spread..... really good insight.


"you're spreading out that drop of spore solution over a bigger surface area and also spreading out whatever is in that drop. Hopefully if you spread the drop out enough, you'll get some mycelium to grow apart from anything else"  SMART. WILL DO!!! Purchased some inoculation loops and will report back with good news hopefully.


--------------------
UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


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Offlinemetalcore_dad
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Re: [FRUSTRATED] Are my syringes contaminated ? What is going on ? [Re: feldman114]
    #26506403 - 02/27/20 12:12 PM (3 years, 10 months ago)

I did microscopy --- unless it was early mold mycelium bc a day later it went from healthy looking to STINKY infested powerdery mold.

Check the microscope images. First 2 were from a plate that I THOUGHT was healthy mycelium but then the next day everything turned to mold. I'm thinking that these mycelium are from a mold mycelium. I'm going to retry this with the same syringes and do the streak method and see if I can isolate something.

Also would very easily believe bunk spore syringes not growing myc at all. Hasn't happened to me after 20+ jars.

Thank you for the insight


--------------------
UNPLUG YA CONTROLLER DOG OOOOOOOOOOOOOOOOOOOOOH FORFEIT MY DUDE OOOOOOOOOOOOOOOOOOOOOOOOOOOH HE FORFEITS !!! MANGO MANGO MANGO MANGO !!


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