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HisDivineShadow
Stranger

Registered: 11/14/08
Posts: 15
Last seen: 14 years, 10 months
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Open air inoculation w self-sealing injection port - no glove box or hood needed?
#9310996 - 11/25/08 01:59 PM (15 years, 2 months ago) |
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Hey everyone,
I'm writing a story in which fungus cultivation plays a key role so I'm here to research the techniques involved. I've read many forum pages already and done a lot of searching, but I haven't been able to find a definitive answer to this question, so pardon me if it gets asked a lot.
Agar described the "optimal lids" for grain inoculation here:
http://www.shroomery.org/forums/showflat.php/Number/6339502#6339502
If you were to use the spawn lids described above, would you need a flow hood or glove box to inoculate grains? It seems to me if you keep your syringe well sterilized, wrapping the needle in an alcohol-soaked rag, you could inject through the rubber stopper to inoculate your jars with minimal risk of contamination. If you inject through an alcohol-soaked paper towel and then through the stopper, shouldn't that destroy or rub away most all fungus spores? Particularly if you inoculate with an LC for quick colonization.
Let me know if I'm wrong about this; thanks for any info you can offer.
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J3illy
Trainee

Registered: 10/18/08
Posts: 3,344
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: HisDivineShadow]
#9311051 - 11/25/08 02:08 PM (15 years, 2 months ago) |
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Well I'll tell you this from experience.
Had 2 syringes and did 2 open-air inoculations. Sprayed oust and sterilized area w/ bleach, and had sterile gloves on cleaned w/ alc. Cleaned ports w/ alc. wipe, which I left on the port. Needles were brand-new, and as I slid the cap off, I slid an alc. wipe over it - so open air contact was extremely brief/ minimal. Both inoculations were THROUGH the alc. wipe, and resulted in germination of spores - but 1 bag got trich right at the inoc. point, which came from the needle.
I did another inoculation w/ that same needle 2 wks later, and flame-sterilized, and did the same exact routine - that bag did NOT get trich.
So my point is, even w/ my utmost care - a trich spore managed to somehow land on that needle in that brief amount of time. I know that it wasn't INSIDE the needle - or else the 2nd bag would have gotten it too. However safe you are - open air will always have some risk involved.
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Typhus
ALGUAALGUALGUALGUALGUALGUAL




Registered: 09/30/08
Posts: 65
Loc: Minnesnota
Last seen: 9 years, 6 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: HisDivineShadow]
#9311157 - 11/25/08 02:27 PM (15 years, 2 months ago) |
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I do open air innocs all the time, all i do is spray lysol, wipe work area/jars with alcohol, wear gloves and flame needle/wipe with alcohol before i inject through normal micropore tape, then put another bit of tape over the hole when im done. I have had 100% success rate doing so, but that is because i have close to no mold (none visible, must be some somewere) were i do it, i run a dehumidifier regularly. I would never advise anyone else to not use a flow hood/glove box, but it works for me.
--------------------
ARE YOU JENNA JAMESON BECAUSE I AM SERIOUSLY STIFF
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seven
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Registered: 06/09/08
Posts: 1,478
Loc: north carolina
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: Typhus]
#9311363 - 11/25/08 03:08 PM (15 years, 2 months ago) |
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you will have more consistant success and waste less material over time using a still air glovebox. period,
-------------------- grind
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iMtRiPpIn
Lookin' fer Luv



Registered: 06/04/08
Posts: 530
Last seen: 11 years, 4 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: Typhus]
#9311375 - 11/25/08 03:10 PM (15 years, 2 months ago) |
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Bulding a still air box is simple, cheap, and effective for minimizing risks associated with open air inoculation. Here is one that can be built in ten minutes for about $17.00. It utilzes a ten gallon fish tank, some cardboard, and some tape. If you can find a ten gallon fish tank at a yard sale or thrift shop, you could probably do this for five or six bucks.

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Edited by iMtRiPpIn (11/25/08 03:13 PM)
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Farmer Joe
HAAAAY!

Registered: 09/24/08
Posts: 44
Last seen: 12 years, 6 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: iMtRiPpIn]
#9311394 - 11/25/08 03:11 PM (15 years, 2 months ago) |
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I just built a glove box out of a cardboard box. I did 7 grain to grain transfers and not a single contamination!
-------------------- Just going with the flow of life....
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dead
grateful



Registered: 08/28/08
Posts: 2,761
Loc: North pole
Last seen: 9 years, 3 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: Farmer Joe]
#9312011 - 11/25/08 04:37 PM (15 years, 2 months ago) |
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-------------------- "The third eye. You spend years doing everything you can to open it and then the damn thing opens and your friends laugh at you when you tell them you can see their souls behind their eyes burning like rainbows." Links:
Nibin's Guide for Noobs
some easy teks on bulk & grain prep. (my journal)
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J3illy
Trainee

Registered: 10/18/08
Posts: 3,344
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: dead]
#9312081 - 11/25/08 04:46 PM (15 years, 2 months ago) |
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Menace gave me an idea for THE MOST simple glovebox ever. Get a clear aquarium, and put it upside down on the edge of a table, but have it a few inches of the edge, so you can slide your hands up into it. There is nothing to make or do or anything like that - and you will have still air. Here's a pic
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fizzbin
Stranger


Registered: 10/09/08
Posts: 64
Loc: Oregon
Last seen: 9 years, 2 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: J3illy]
#9312449 - 11/25/08 05:50 PM (15 years, 2 months ago) |
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I'm not sure how you work with LCs aside from removing the cap.
I injected 10 jars with a LC that was just sitting in a Miracle Whip jar on my bathroom floor, using a spent 10cc syringe, and got no contamination.
> J3illy
Your problem may of been when you put the needle onto the syringe. There's air from the room that gets stuck in there. I'm assuming that giving it a quick squirt onto a towel removes that.
-------------------- [quote]It is far better to grasp the universe as it really is than to persist in delusion, however satisfying and reassuring.[/quote] [cite]-- Carl Sagan[/cite]
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RogerRabbit
Bans for Pleasure



Registered: 03/26/03
Posts: 42,214
Loc: Seattle
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: fizzbin]
#9312471 - 11/25/08 05:56 PM (15 years, 2 months ago) |
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I would never recommend 'open air' anything related to inoculations of grains, petri dishes, or even pf cakes. It's just too easy to build a simple still air box as shown in more than one post above. RR
-------------------- Download Let's Grow Mushrooms semper in excretia sumus solim profundum variat "I've never had a failed experiment. I've only discovered 10,000 methods which do not work." Thomas Edison
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J3illy
Trainee

Registered: 10/18/08
Posts: 3,344
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: fizzbin]
#9312492 - 11/25/08 06:01 PM (15 years, 2 months ago) |
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Quote:
fizzbin said: > J3illy
Your problem may of been when you put the needle onto the syringe. There's air from the room that gets stuck in there.
Ya kno, that's exactly what I thought. But if this is what happened, then I would think the 2nd bag I knocked w/ that syringe 2 wks later also would have gotten trich, cuz the contam would have been inside w/ the liquid. But after flaming the needle and knocking another bag - that 1 is doing great. No trich or any contams at all. So however unlikely it is - it really seems that a spore somehow landed on the needle itself. The fact the trich grew RIGHT at the inoc. point too, is pretty damning evidence, lol.
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Stimpy913
Happy Happy Joy Joy

Registered: 11/05/08
Posts: 765
Last seen: 14 years, 10 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: J3illy]
#9312721 - 11/25/08 06:47 PM (15 years, 2 months ago) |
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Takes 20 seconds to cut some arm holes out of cardboard with a steak knife, another 10 seconds to put some see-thru wrap over the top and tape it shut. The point of a glovebox isn't even to be sterile. It's just to make a small closed off environment, where air currents cant push contam spores about. Simply opening a door will put contam spores flying all hazardly across a room. I'd use bleachwater spray on the glovebox, because alcohal is flamable.
-------------------- How to talk to cops
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dothedew69
Relearning



Registered: 08/27/08
Posts: 624
Last seen: 3 years, 3 months
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: Stimpy913]
#9313634 - 11/25/08 09:02 PM (15 years, 2 months ago) |
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Glovebox with arms like mine is just way too clumsy and a PIA. I hate being restricted so I just Oust (made for air unless you use the Lysol made for air), face mask, gloves, good anti-bacterial cleaning, alcohol whipe needle, flame sterilize, inject. 1 Contam so far... and that was in a still air glove box! So out of 50 jars nothing yet. I am not saying this is foolproof because I am not retarded. You would be best using a still air box or a flowhood. Until I can pony up for a flowhood I won't bother with a glovebox. I would rather make 20 jars and lose 3-6 just for the convenience factor. But of all the jars I had the only contam I had was from a glovebox.
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billyboy36
Stranger

Registered: 05/04/08
Posts: 352
Loc: Canada
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: HisDivineShadow]
#9314447 - 11/25/08 11:24 PM (15 years, 2 months ago) |
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glove boxes aren't guaranteed either. If you ask me, they're a crutch when doing simple grain jar inoculation. People can get sloppy because they think the glove box is going to save their ass. They also restrict movement. Now, I'm speaking strictly of simple grain jar inoculations. G2G's and cloning etc. are better off in a still air box or in front of a flowhood.
oust and turn down or off the air con or furnace and all fans a half hour prior to inoculation.
Simply soak a paper towel with iso alcohol and rub it vigorously on the port. Then LEAVE it on there. Starting at the back of the needle, flame it red hot. Do not let the needle touch anything or wipe it down with another alcohol rag. Bring the needle to within an inch of the port, wipe the port with the alky towel one last time and squeeze a few drops out of the syringe to cool the needle. Then immediately inject just enough of the needle (1/4 or so for grain jars) into the port and inoculate. There's no reason to put more needle into the jar than you need. And since you brought the needle so close to the port before you cooled it down, the amount of time that the needle is exposed to air is less than a second.
It's never failed me and it's not a bad practice to get into even if you are using a glove box or flow hood. My .02
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Rickster
El Conquistador



Registered: 02/06/06
Posts: 659
Loc: Babylon
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Re: Open air inoculation w self-sealing injection port - no glove box or hood needed? [Re: billyboy36]
#9314547 - 11/25/08 11:50 PM (15 years, 2 months ago) |
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a clear plastic trashbag dose the trick for inoculations and g2g transfers for me.
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