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 camplo
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 Inoculating jars via lifting the lid.
#8595966 - 07/04/08 08:37 PM (3 months, 7 days ago) |
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Just wondering if anyone had any advice on the topic. I introduced a colonized p[late of agar into a jar of rye via lifting the lid and pouring the agar chopped as best I could inside its jar.
Jar went green like 6 days later!
So I knew I Skimped on PC time. i think I only hit like 70 minutes but it was only 100 grams of rye I mean come on. Maybe I had a lid breach cause I think my filter might of been funny but its a big might.
So this time I did a generic lid setup of mine (one hole one tyvek one medical tape) simple and time proven. When I opened the jar (in the glovebox of course) the jar had a slight vacuum cause it was cooler in my glovebox than where it was sitting. I inoculated with only .5cc of spore solution (yeah Im a cheapy) and its been 3 days and no noticeable growth. Im sure I'll see something by like day 6. Hopefully no green though.
Edited by camplo (07/08/08 04:33 PM)
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 tripsis
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Loc: Gondwana
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Re: Inoculating jars via lifting the lid. [Re: camplo]
#8596157 - 07/04/08 10:32 PM (3 months, 7 days ago) |
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Lifting the lid with an agar wedge is the only way to inoculate. Rather than tipping the plate over the mouth of the jar, carefully pick up the wedges with a scalpel and drop them into the jar. Do it as quickly as possible and try not to lift anything other than the wedge above the jar opening.
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Since we depend on an abundance of functioning ecosystems to cleanse our water, enrich our soil and manufacture the very air we breathe, biodiversity is clearly not an inheritance to be discarded carelessly - Edward O. Wilson 1992
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happyday
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Re: Inoculating jars via lifting the lid. [Re: camplo]
#8597324 - 07/05/08 12:39 PM (3 months, 6 days ago) |
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You could also utilize a semi-sealed (filtered) or sealed container for liquid culture in conjunction with a spawn bag with some type of mechanism for injection site, i.e. even a dab of silicone (teks are listed here).
70 minutes is more than enough time for jars smaller than two gallons (@15psi). Green molds do not necessarily mean you "skimped on pc time". They could have creeped in anytime during procedure.
Anywhere is going to be "cooler" than the autoclave. Not just your glovebox.
Use a greater quantity of spore solution than .5 cc. You'll have better results.
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camplo
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Re: Inoculating jars via lifting the lid. [Re: happyday]
#8608958 - 07/08/08 04:42 PM (3 months, 3 days ago) |
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Yeah but I'm a conservative, I rather I use the least amount possible. 1/2cc of inoculated should have been enough to produce germination. That last jar I did I have not received any growth. I'm sure it has something to do with the small amount of inoculate, it just seems like .5ccs should be enough when I use so much less to get germination on agar. I'm just just glad it did not turn green. I think maybe I didn't clean my glovebox and work area enough last time? Probably... You are right about how the temperature difference creates a vacuum inside of the jar, an effect that increases with increased contrast of temperatures... it just sounds better when I say it. I'm sorry, I don't mean to pick I'm just sayin "anywhere could be cooler than my PC"? Duh? I think you are referring to about how I said something about when I opened the jar and it had a vacuum right? It had been out of the PC for many many an hour before I attempted to open its lid.
The vacuum deal was my fault because I had the jar previously in the incubator which probably was a few degrees over room temp. Doesn't matter though cause the damn jar never showed signs of life.
Edited by camplo (07/08/08 05:13 PM)
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camplo
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Re: Inoculating jars via lifting the lid. [Re: camplo]
#8609357 - 07/08/08 06:29 PM (3 months, 3 days ago) |
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Tripsis this is directed towards you. I was just thinking that the only reason why I thought to inoculate spore to grain was because I was thinking that lets say I just want to inoculate just one jar of rye I would only germinate the strains located on that section of agar. I want all of them that germinate, never know might leave out that "super strain" and never get to clone it. So Id be looking to slice up the whole thing to put in one jar. I kinda tried this once but I did not work out for me in the end. Maybe my agar was too solid but I was not able to cut up the agar so easily, got maybe 4 pieces separated then was like fuck man I don't like having this jar open for so long lets get this over with and just dumped those 4 chunks into the jar. Ended up green stuff in the end but I think maybe my loading procedure and general area cleanliness might not of been up to par. Getting to cocky with me fancy glovebox sorta thing and not following the rules, not to mention what you said about the more better tighter procedure.
Edited by camplo (07/08/08 06:31 PM)
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RogerRabbit
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Re: Inoculating jars via lifting the lid. [Re: camplo]
#8611929 - 07/09/08 07:24 AM (3 months, 2 days ago) |
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There shouldn't have been a vacuum in the jar. Within minutes, the tyvek should have equalized pressure. Perhaps your tape isn't breathable. Something else was wrong, but definitely don't pour from a petri dish into a jar of grains. Never have a petri dish or grain jar open for over five seconds. Inoculate quickly with a wedge of agar, not the whole dish.
RR
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camplo
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Re: Inoculating jars via lifting the lid. [Re: RogerRabbit]
#8617951 - 07/10/08 03:34 PM (3 months, 1 day ago) |
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FRom the best I can tell its the same medical tape that I see everyone else using by 3m. Its just Tape holding a patch of tyvek of a hole thats about 1/4th of an inch. Yeah it does seem that it shouldnt have done that. Doesnt matter to me now cause it did not cause anything bad to get sucked in right now Im like hey! Wheres the germination! Doesnt smell funny.....what gives.....1/2ccs..... is it...e......nough?
The 5 second rule you speak of applies to the inside of the glovebox too?
Edited by camplo (07/10/08 03:40 PM)
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camplo
Freedom!
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Re: Inoculating jars via lifting the lid. [Re: camplo]
#8617962 - 07/10/08 03:36 PM (3 months, 1 day ago) |
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OK fuck it I'm gonna try the same jar again. First I'll weigh it and figure its water content then as long its not too low I'll use 2ccs this time. Anyone know if prints from ralphsters have shipping included or not? The order form is a little off to me...
Edited by camplo (07/10/08 03:36 PM)
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RogerRabbit
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Re: Inoculating jars via lifting the lid. [Re: camplo]
#8624525 - 07/12/08 07:43 AM (2 months, 30 days ago) |
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I follow the five second rule even with a flowhood.
RR
--------------------
www.mushroomvideos.com
"I wouldn't want to belong to any club that would have me as a member".
Mark Twain, Woody Allen, Groucho Marx, and anyone else who wishes to claim credit for the quote.
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