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damn trich is back


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#21 I_am_me

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Posted 24 September 2006 - 06:22 PM

Heh well we'll just have to agree to disagree. ;)
Although one last thing about fractional sterilization. It was intruduced by John Tyndall and used to show the existence of heat-resistant bacterial spores, which is why it specifically refers to more than one sterilization with an incubation time inbetween. I think he published his method in 1877.
http://en.wikipedia....ki/John_Tyndall
http://dictionary.la...alk.com/Tyndall
That is why it is also called Tyndallization. Heh, I read a lot about this before I purchased an All American Pc to use for popcorn.
Sorry to get off topic teletrue. I think you should try your hand at making your own syringes. Do you have a variety of syringes to choose from this time around?

#22 teletrue

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Posted 24 September 2006 - 07:08 PM

So, before I begin, is there any advice about something I may be missing during the sterilization process? Throw all your tips out here, and I'm going to do my damndest to make a batch of clean jars.

#23 srgtm1a

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Posted 24 September 2006 - 07:19 PM

So, before I begin, is there any advice about something I may be missing during the sterilization process? Throw all your tips out here, and I'm going to do my damndest to make a batch of clean jars.



Just follow the procedures step by step, even if you've done it a bunch b4....sometimes something as small as leaving a fan on could do you in....and is easily forgotten.

Just make sure your work surface is clean, rub down with rubbing alcohol etc, and to take extra percaution...what I do with syringes is swab with rubbing alcohol, flame, swab again, let dry, then inoculate. It's overkill, but it does wonders in the way of being safe.

#24 teletrue

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Posted 24 September 2006 - 07:21 PM

what I do with syringes is swab with rubbing alcohol, flame, swab again, let dry, then inoculate. It's overkill, but it does wonders in the way of being safe.


Damn..that's what I do too. :-/

#25 teletrue

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Posted 24 September 2006 - 08:07 PM

ok. So. I loaded the jars as clean as possible, and used hand sanitizer through the whole process. I've included pictures of my procedure...

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#26 Guest_CoyoteMesc_*

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Posted 24 September 2006 - 08:59 PM

where did Lazlo go after that good analogy..
LMFAO

#27 teletrue

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Posted 25 September 2006 - 12:05 AM

So, after the jar came out of the pot, and was allowed to cool somewhat, I took rubbing alcohol, wiped the whole lid off and put two dabs of silicone sealant on the top foil layer, leaving the jar lid on. I think that the probability of contamination will be reduced if I use a self-healing port to innoculate. I'm going to SSS tomorrow morning, and then noc it up.

So, now for open air innoc advice...

#28 apokalypse

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Posted 25 September 2006 - 01:45 AM

Is the silicon on the jar lid or the tinfoil covering the jar lid? I hope it's on the jar lid itself and not the foil. Do you take your tinfoil off before innoculation and leave it off?

#29 teletrue

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Posted 25 September 2006 - 08:26 AM

it's mostly on the foil, but some is on the jar lid ring..

#30 BuckarooBanzai

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Posted 25 September 2006 - 09:24 AM

Don't forget, your jars need some fresh air to grow to completion.

If you seal the jar up tight with silicone, it will stall on you.

#31 Cryingblackoil

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Posted 29 September 2006 - 09:12 AM

Still air. Thats a very important thing. Turn off the A/C a long time before you plan on starting. Maybe an hour or so. Does wonders.

#32 kaneda182

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Posted 29 September 2006 - 05:59 PM

:greenboun:greenboun:greenboun:greenboun:greenboun:greenboun:greenboun:greenboun:greenboun

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#33 teletrue

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Posted 12 October 2006 - 02:28 PM

So, the next day after the jar cooled, I went into my bleach bombed bathroom, and despite the fumes (about killing me), I took my time and carefully (and very sterily) innoculated with an EQ spore syringe. Wiping innoc. points with alcohol, wiping down the needle with alcohol, then flaming, then wiping with alcohol again before innoculating.

Almost a month later, and I have an almost pink/red thing going on...tell me what you think...I really cannot believe that this contamination is coming from anywhere but the syringe. I know it's not trich. But, it came from a different syringe from the same supplier.

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#34 teletrue

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Posted 23 March 2007 - 02:10 PM

So,

After getting myself a pressure cooker, and PC'ing my jars correctly, I noc'd them up using three of the four syringes I got from sporelab last fall. The other jar was noc'd up using a syringe I made with a spore print of the HillBilly strain.

Well. 4 days later, there's good white growth in only one of the jars - the one from the spore syringe I made.

The other three have the same damn trich that I was getting at the original posting of the message, leading me to believe I recieved bad syringes.

As previously stated, I had always used steam sterilization to sterilize my half pints and have NEVER EVER had problems like this.
What was different this time, however, was the trial of a new spore vendor (sporelab).

I'm not trying to bash sporelab, as I know a lot of others have had great success with their prints and syringes. I'm just telling my story. A lot of people told me that it was my method of sterilization - which again, has never given me problems like this - so I bought a pressure cooker and have used it - correctly I might add.

If anybody can think of ANYTHING other than bad syringes which would have given me these results, PLEASE let me know. Prior to this, I have used another one of mycotopia's sponsors as a spore supplier and have had great results. I think I'm going to go back.
By the way, that perticular grow was 36 half-pints. ALL STEAMED for an hour and a half. They were noc'd up in a glovebox (as these were..) and I had ZERO come up contaminated. Now, with the only difference being a new supplier, every single jar has come up contaminated.

Something smells.

#35 Lazlo

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Posted 23 March 2007 - 02:17 PM

In the pictures above, i'd have to think the spore syringe was bad. I too steam sterilized all of my pf jars and never had too much of problem with contamination. Unless it was from a bad print I had taken.

#36 teletrue

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Posted 23 March 2007 - 02:22 PM

In the pictures above, i'd have to think the spore syringe was bad. I too steam sterilized all of my pf jars and never had too much of problem with contamination. Unless it was from a bad print I had taken.



THANK YOU THANK YOU THANK YOU.

Finally. Someone believes me.

#37 Lazlo

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Posted 23 March 2007 - 02:40 PM

It's obvious really. You'd really have to screw up royally in order for that to happen. I mean royally. Like dipping the needle in mold spores prior to inoculation. lol

#38 teletrue

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Posted 23 March 2007 - 04:31 PM

Yeah, and I tend to be pretty OCD about cleanliness when it comes to having to have something COMPLETELY sterile. Having the pressure cooker only confirms my beliefs about the syringes I recieved, which were replacements already for....contaminated syringes. *sigh*

#39 dinosaur

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Posted 23 March 2007 - 08:14 PM

I never used a hot pot either, never had problems. I use ISO 99% on everything, boil jars for 1 hour 15 minutes and don't hardly get contams. 1/50

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#40 nrthlndr27

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Posted 24 March 2007 - 01:19 PM

Have you ever run a 'method blank' on your procedure? In analytical labs it's standard operating procedure to run a method blank along with your batch of samples. When looking for volatile organic compounds in the parts per billion range, samples can easily have false positives for common lab solvents (methylene chloride contamination is common) and the way you weed out whether it is actually in the sample or introduced via your procedure is to run a method blank. If, when the method blank is analyzed along with the rest of the samples, has positive detects for any analyte then you know it is in your procedure, not the sample.

The same theory is easily applied to mushroom cultivation, especially now that you have a pressure cooker. Sterilize some syringes of distilled water in your PC. When innoculating your batch of jars save a few to run several experiments. On several, use the sterilized syringes of water and shoot up your jars just like you would with spore solution. On several more leave them untouched. Handle and incubate these jars just like you would your normal jars.

If you have contamination problems in innoculated jars, water jars, and untouched jars, then I would conclude the problem lies in your substrate preparation method.

If you get contamination in the jars with spore solution and jars shot with sterile water, but not in the blank jars, then contamination is being introduced during the syringe procedure.

If your spore solution jars are contaminated, but your sterile water jars and your blank jars show no signs of contamination, then you're working with bad spore syringes.

I was having problems with bacteria contamination and thought it was my liquid culture. I ran some method blank jars and figured out it was my sterilization procedure. Most jars were going bad with bacteria. I was PCing for 75 minutes at 15 psi but was loading my pressure cooker to full capacity. My conclusion was the PC was too full to completely sterilize its contents for the time I was heating. My LC was fine.




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