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raoulduke69
Pretty Cool Guy


Registered: 07/12/20
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Reddish Purple in Agar Plate
#26896082 - 08/23/20 09:49 PM (3 years, 5 months ago) |
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So I've started experimenting with agar using pastywhite's easy agar tek. It has been 15 days since I made my first attempt. One of the containers shows a spot of white surrounded by reddish purple. Researching the site suggests perhaps it's fusarium. I wanted to get others' input as well. Since I didn't use an inoculation loop my first time, and instead just put a single drop of spore solution in, I figure this plate is probably toast.

As always, thanks for any input.
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Rhizomorph
Psychedelic Researcher



Registered: 04/24/20
Posts: 785
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Re: Reddish Purple in Agar Plate [Re: raoulduke69]
#26896088 - 08/23/20 09:54 PM (3 years, 5 months ago) |
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Definitely toast.
If you want to transfer and try to clean it up this is an option. But you're probably better off inoculating a fresh plate.
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Rhizomorph
Psychedelic Researcher



Registered: 04/24/20
Posts: 785
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Re: Reddish Purple in Agar Plate [Re: Rhizomorph]
#26896090 - 08/23/20 09:58 PM (3 years, 5 months ago) |
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Also chances are it is contamination in the spore syringe/solution assuming you followed pasty's tek to a T.
Spore syringes tend to be a bit hit or miss since they're virtually never contaminant free. They generally just have a low enough contaminant:spore ratio that you can usually get away with making BRF cakes & whatnot.
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dirtybirdx
getting better

Registered: 07/28/20
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Re: Reddish Purple in Agar Plate [Re: Rhizomorph]
#26896125 - 08/23/20 10:34 PM (3 years, 5 months ago) |
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Quote:
Rhizomorph said: Also chances are it is contamination in the spore syringe/solution assuming you followed pasty's tek to a T.
Spore syringes tend to be a bit hit or miss since they're virtually never contaminant free. They generally just have a low enough contaminant:spore ratio that you can usually get away with making BRF cakes & whatnot.
I keep seeing this repeated over and over. I understand the accepted consensus that spore syringes are generally dirty. But this is probably only accurate if you’re talking about the entire population of spore syringes.
Most people buy spore syringes from reputable vendors, and in this case, they’re usually good. Most people aren’t using ones they made themselves or were made by other home culters.
So why am I commenting? Because I’ve never experienced a ‘bad’ syringe.
Sure, I’ve only ordered from reputable vendors as mentioned, but I’ve also purchased close to a dozen syringes over the years (not a huge sample, I know) and have never had one that wasn’t usable due to contam.
Take ‘syringes are dirty’ with a grain of salt, and buy from reputable vendors.
Advice like this might lead the guy to toss his syringe and buy another one when it’s actually probably his sterile tek. It’s more likely that a random contam made it into his jar during inoculation.
And even if that’s not what actually happened - he will learn more by trying to tighten up his sterile tek and still come to the conclusion that his syringe is bad rather than spending more money on a syringe and potentially throwing away usable spores. It’s never happened to me, but he would learn a ton that way.
Spore Syringe Lives Matter
-------------------- Need help?

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Rhizomorph
Psychedelic Researcher



Registered: 04/24/20
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Re: Reddish Purple in Agar Plate [Re: dirtybirdx]
#26896304 - 08/24/20 01:56 AM (3 years, 5 months ago) |
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Quote:
dirtybirdx said: Advice like this might lead the guy to toss his syringe and buy another one when it’s actually probably his sterile tek.
What advice? I just mentioned what I believe to be the likely source of contamination. I never said anything about tossing syringes or that syringes shouldn't be used. I agree they have their place - all cultures must come from somewhere right?
Also you may not have had bad syringes but I've seen many people who have. Based on the information in the OP it is really a shot in the dark since the OP didn't cover whether the syringe came from a reputable source or whether they had proper sterile technique. As I said I'm just mentioning what I believe to be the most likely source, but we don't have enough data to accurately measure that for sure.
I think improving sterile technique is a great idea but so is understanding that there are variable potential contamination points including both sterile technique & spore syringes. I think the OP could benefit from both. Just my
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raoulduke69
Pretty Cool Guy


Registered: 07/12/20
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Re: Reddish Purple in Agar Plate [Re: Rhizomorph]
#26896465 - 08/24/20 06:26 AM (3 years, 5 months ago) |
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I ordered from a reputable vendor.
I've struggled with sterilization with both BRF and agar. At this point, I'm using rubber gloves and tyvek sleeves sprayed down with isopropyl, a mask, a SAB that I mist with a water/soap mixture and place a towel underneath, and I flame the syringe needle, of course. One nuisance that I'm encountering is that where I currently have to operate, there is a dog in the house, whose hair gets on absolutely anything and everything. I even find stray hairs on my SAB after it has been in storage for a while. So, I'm thinking maybe I ought to clean the SAB with something stronger than water in a spray bottle with a drop of soap.
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Munchauzen


Registered: 06/22/11
Posts: 14,342
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Re: Reddish Purple in Agar Plate [Re: raoulduke69]
#26896474 - 08/24/20 06:48 AM (3 years, 5 months ago) |
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most likely its not the cleanliness of your house, just technique / bad luck. I would just toss it and move on with your life, not something worth making changes over. do more plates.
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Gan
Wielder of Narya



Registered: 08/26/19
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Re: Reddish Purple in Agar Plate [Re: raoulduke69]
#26896476 - 08/24/20 06:49 AM (3 years, 5 months ago) |
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Clean it with some 70% IPA beforehand. Then spray it down. Are you streaking your spores across the plate or just placing a drop in the center?
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raoulduke69
Pretty Cool Guy


Registered: 07/12/20
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Re: Reddish Purple in Agar Plate [Re: Gan]
#26897808 - 08/24/20 08:25 PM (3 years, 5 months ago) |
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The idea of using ISA first and then spraying had occurred to me. Will definitely do that next time.
This first time I just used a drop. Next time I will streak. I actually did attempt to streak a plate. I made an inoculation loop, but then failed to use it correctly. I forgot you're supposed to cool the loop in the agar. Is it correct that by sticking the loop briefly in the agar, you get some residual agar attached to the loop, which then allows the spore drops to attach to the loop and then drip onto the plate?
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Gan
Wielder of Narya



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Re: Reddish Purple in Agar Plate [Re: raoulduke69]
#26897821 - 08/24/20 08:33 PM (3 years, 5 months ago) |
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Yeah so even using spore solution, you can do it a few different ways.
You can cool it off in the agar, then either drop a drop of spore solution on the loop itself and then streak. Or put a drop somewhere on the plate, dip your loop in it, and start your streak pattern from wherever the initial drop is.
Alternatively, you can flame your loop, drop 1-2 drops of spore solution on it to cool it off, then 1 more drop and streak. It all depends on what works for you and what you like best. All should work fine with a viable syringe.
Prints are bit easier to go to agar with, but when I first started all my spores were in solution so that's where I started. Works just fine. As long as you dont use more than a drop. Shake it up well beforehand as well
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raoulduke69
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Re: Reddish Purple in Agar Plate [Re: Gan]
#26899979 - 08/26/20 06:17 AM (3 years, 5 months ago) |
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Thank you for this excellent explanation, friend. This will prove quite helpful. When I attempted to streak last time, I just missed the inoculation loop when I attempted to drop spores onto it. Now I know I could've still streaked by dipping the loop into the agar where the drop fell.
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