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Offlinepolaritymind
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LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP
    #26762983 - 06/21/20 12:24 PM (3 years, 7 months ago)

Ok so I used to fuck around with LCs and stopped, now Im back to it. To increase my rates of sucess this time I wanna adapt some more strict controls, like only using an LC once. (They were often fine at first and then contamed). Even though testing an LC on agar, goes against this, but thats also a nex step I'd like to introduce. And also not opening the jars and doing everything through SHIPS/ports. And this brings me to my question, is that even possible?
Is it possible to suck up LC in sufficient amounts through a SHIP/port (with a syringe) without creating a vacuum, and the syringe not moving further? I am using Polyfill Filters that are very tight.

And doesnt using an LC for inculation mess up the moisture content of my grain jars for example? And if you just use so little LC, so that it doesnt, is there really any benefit in terms of "more points of inoculation" compared to just dropping in a wedge of Agar, which is how I've been doing it so far?


Thanks for your help.


Edited by polaritymind (06/22/20 05:11 AM)


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Offlinepolaritymind
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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: polaritymind]
    #26776452 - 06/23/20 10:53 AM (3 years, 7 months ago)

no one?


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InvisiblePastywhyteMDiscord
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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: polaritymind]
    #26776458 - 06/23/20 10:56 AM (3 years, 7 months ago)

If there is no gas exchange in the vessel due to the filter being too tight, you definitely won’t be able to aspirate anything out of it. If you cannot get it out at all I would recommend trying to let it grow a bit extra thick and then pour it like G2G.


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Offlinepolaritymind
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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: Pastywhyte]
    #26777096 - 06/23/20 03:38 PM (3 years, 7 months ago)

And then what about the worry with the moisture content getting too high then?


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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: polaritymind]
    #26777204 - 06/23/20 04:14 PM (3 years, 7 months ago)

Pour less if you don’t want it to be soaked. Let it colonize a bit thicker and it’s easy to do.


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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: Pastywhyte]
    #26778661 - 06/24/20 08:07 AM (3 years, 7 months ago)

I mostly lifted the lid of the jar a little to equalise the pressure so I could then squirt in some spores without having the whole syringe emptied. I never had one contaminate, even when I used a glove box.

If you have a clean syringe that only has a small amount of liquid left then it wouldn't matter if the liquid culture jar sucked all of the remaining contents of the syringe into the jar. I have done it this way before and I made a self healing injection port by using silicone. 

I've also made liquid cultures using a flow hood but that makes sterile conditions a lot easier and if you have a flow hood then it's easier to just do a bunch of grain to grain transfers to make your spores or mycelium go further.

Using a liquid culture syringe to inoculate your grain shouldn't make the grain soggy unless you're aspirating too much liquid in a small container. Just make sure your grain isn't already too wet or soggy when you fill your jars or bags.


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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: migraineur]
    #26778669 - 06/24/20 08:09 AM (3 years, 7 months ago)

Spores to LC? Lifted the lid to equalize the pressure? No contams ever? Glovebox?
Sounds like bullshit to me.


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Offlinemigraineur
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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: Pastywhyte]
    #26778701 - 06/24/20 08:26 AM (3 years, 7 months ago)

No joke. I was extra thorough with my cleaning and sterile technique but perhaps it was that combined with luck. Either way I never had one contaminate. I only ever had one not work at all and I think it was because it ran out of oxygen as there was no filter for gas exchange.

I had gotten dirty syringes off others before but I always tested those on one or two sacrificial test jars before using them for anything more or else.


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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: migraineur]
    #26778706 - 06/24/20 08:29 AM (3 years, 7 months ago)

If you never had a contam you simply are not growing much. Even commercial farms and spawn labs get contams eventually, and that’s in ISO level 6 clean rooms and crazy stringent procedures. If you never had a contam shooting biological material collected in less than axenic conditions, you simply haven’t done it much. I can’t even imagine a person doing 50 grows in this manner before it eventually caught up with them.


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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: Pastywhyte] * 1
    #26778750 - 06/24/20 08:52 AM (3 years, 7 months ago)

Pretty much. Labs that grow yeast for breweries get hit with contaminants from time to time and they're doing everything in clean rooms.
Also this guy has probably had plenty of contamination that he just didn't recognize


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Offlinemigraineur
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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: polaritymind]
    #26778779 - 06/24/20 09:10 AM (3 years, 7 months ago)

I've done plenty of grows but I eventually moved on from liquid cultures when I could easily do grain to grain with my flowhood.

I definitely had contamination when I was doing grain to grain in a glove box and agar work. I guess I just got lucky with the liquid cultures I made previously.


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Re: LC Questions - Moisture Content and neg. pressure in syringe when sucking out through SHIP [Re: Pastywhyte]
    #27236881 - 03/04/21 11:55 AM (2 years, 10 months ago)

Quote:

Pastywhyte said:
Spores to LC? Lifted the lid to equalize the pressure? No contams ever? Glovebox?
Sounds like bullshit to me.




Just thought I'd let you know that I had an untested LC in the fridge that I went to throw away due to age. I looked into the jar and saw what looked like obvious contamination so it eventually happened to me. The first thing I thought of what this old thread.

The guy whom I got my syringes from doesn't exactly have the best sterile technique though.


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