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Offlinejomanda1990
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Re: The Biopsy Method: A different way of doing LC. [Re: Mr. Funguy]
    #26520687 - 03/06/20 04:02 PM (1 year, 7 months ago)

Bumping this method and I'd like to ask something.
I got 2 jars with grain soak water diluted to 50% and transferred a biopsy from a clone on agar. These jars are showing (after 4 days) some mycelium puffballs in the bottom of the jar, so I guess they are fine.

I also had a clean agar plate with MS grow and some hyphal knots. I poked one of them and transferred this biopsy to a jar (also with a grain water LC broth). It's been 4 days and now there's like.. a ghost in there? It looks like a pair of napkins floating in the jar. WTH is that? :confused:



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Invisiblewoofwoof
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Re: The Biopsy Method: A different way of doing LC. [Re: Josex]
    #26523185 - 03/08/20 04:01 AM (1 year, 7 months ago)

I know the tek says to PC the syringes, is this because boiling water is not truly sterilized?

I am about to do this method, but wondering if I can boil the syringe water and use that. It works for spores, but spores are dirty any ways. I just want to be sure, and my gut tells me to follow the tek, but was curious


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Invisiblewoofwoof
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Re: The Biopsy Method: A different way of doing LC. [Re: woofwoof]
    #26523332 - 03/08/20 06:25 AM (1 year, 7 months ago)

Quote:

woofwoof said:
I know the tek says to PC the syringes, is this because boiling water is not truly sterilized?

I am about to do this method, but wondering if I can boil the syringe water and use that. It works for spores, but spores are dirty any ways. I just want to be sure, and my gut tells me to follow the tek, but was curious



Quote:

woofwoof said:
I know the tek says to PC the syringes, is this because boiling water is not truly sterilized?

I am about to do this method, but wondering if I can boil the syringe water and use that. It works for spores, but spores are dirty any ways. I just want to be sure, and my gut tells me to follow the tek, but was curious




I ended up sterilizing it with a PC for 15 min @ 15 PSI. I did 5 plates of blank agar and poked my rusty whyte agar plate. I only have 18 gauge needle and poked thru till i hit the bottom of the dish, then squirted a drop or 2 unto blank agar.

Lets see what happens!!


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Offlinebongoman
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Re: The Biopsy Method: A different way of doing LC. [Re: woofwoof]
    #26528035 - 03/10/20 09:23 PM (1 year, 7 months ago)

How small can I make an LC? I don't have the need for lots of LC so was wondering what's the smallest volume of broth I can make up in practical terms?

Would 100ml be too low a volume?


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OfflineTweeq
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Re: The Biopsy Method: A different way of doing LC. [Re: bongoman]
    #26528398 - 03/11/20 03:19 AM (1 year, 7 months ago)

100ml is fine Ive done even smaller batches


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OfflineZillard
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Re: The Biopsy Method: A different way of doing LC. [Re: Tweeq]
    #26530281 - 03/12/20 06:58 AM (1 year, 7 months ago)

Awesome write up. Thanks.


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OfflineThrill
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Re: The Biopsy Method: A different way of doing LC. [Re: Zillard]
    #26605984 - 04/16/20 04:29 PM (1 year, 5 months ago)

Bumping this cause it looks interesting and I got a question.

Anyone tried this through an injection port instead of shooting through a filter or opening and squirting it in? Or does pushing through the injection port rub off all the tissue cells?

Making a few jars of broth and thought Id give this a shot, see how it works vs some other methods that have popped up in the few years since I been away from here.


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Offlinebongoman
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Re: The Biopsy Method: A different way of doing LC. [Re: Thrill]
    #26606526 - 04/16/20 09:29 PM (1 year, 5 months ago)

Another question about this tek.

Josex says this is "a pour tek — don't aspirate the LC from the jar".

I wonder why not? What if I use a modded lid with an SHIP and a syringe filter, why can't I aspirate to a 60ml syringe with a 16G needle?


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Invisiblewoofwoof
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Re: The Biopsy Method: A different way of doing LC. [Re: bongoman]
    #26607090 - 04/17/20 03:19 AM (1 year, 5 months ago)

I would not do a poke with needle then inject thru a SHIP, the material inside the needle (the myc) may contact the ship and contam it. Don't do it, I have tried and failed here.


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OfflineThrill
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Re: The Biopsy Method: A different way of doing LC. [Re: woofwoof]
    #26607524 - 04/17/20 10:51 AM (1 year, 5 months ago)

I had that thought too, since you cant flame sterilize it once you pull the biopsy. Ill just crack the jar open and squirt it in.


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Offlinebongoman
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Re: The Biopsy Method: A different way of doing LC. [Re: Thrill]
    #26620497 - 04/22/20 06:34 PM (1 year, 5 months ago)

Wow, it really works.

Mexicana LC 11 days after the poke:



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OfflineDoc9151M
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Re: The Biopsy Method: A different way of doing LC. [Re: bongoman]
    #26624387 - 04/24/20 12:31 PM (1 year, 5 months ago)

I don't know if this has been asked and answered, but when it comes to covering the inoculation port, has anyone used hi temp rtv to Make a self sealing port?

edit: never mind,  I  remember yall call it a SHIP


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Psilocybe cubensis data collection thread. please help with this project if you hunt wild cubensis.
https://www.shroomery.org/forums/showflat.php?Cat=0&Number=26513593&page=0&vc=1#26513593


Edited by Doc9151 (04/24/20 12:34 PM)


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OfflineDoc9151M
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Re: The Biopsy Method: A different way of doing LC. [Re: Doc9151]
    #26636550 - 04/29/20 12:56 PM (1 year, 5 months ago)

Josex, I was just reading the LME LC recipe and you stated that you like to use 2g lme vs 1g to get extra nutes. Have you ever tried adding yeast to the recipe instead of the extra lme? I use yeast in my pasty plates occasionally when I want extra nutrients or use a tea made from hpoo to make up the agar instead of plain tap water. 

This tek of yours is my favorite so far, I'm about to make some now.

edit: was bitten by the autocorrect bug.


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Psilocybe cubensis data collection thread. please help with this project if you hunt wild cubensis.
https://www.shroomery.org/forums/showflat.php?Cat=0&Number=26513593&page=0&vc=1#26513593


Edited by Doc9151 (04/29/20 12:58 PM)


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InvisibleJosex
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Re: The Biopsy Method: A different way of doing LC. [Re: Doc9151]
    #26636675 - 04/29/20 01:50 PM (1 year, 5 months ago)

No, I like it easy and lazy if I can so I always use plain LME, works like a charm and if it aint broke...
I've seen no need to add more than one ingredient or go out of your way trying to find a super badass and balanced broth formula to keep myc happy, the LC is just a mean to an end, not the end itself. You just want some healthy myc in there to noc your projects with and LME by itself does a fantastic job, as well as any other grain based broth.


Edited by Josex (04/29/20 01:57 PM)


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OfflineDoc9151M
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Re: The Biopsy Method: A different way of doing LC. [Re: Josex]
    #26636707 - 04/29/20 02:01 PM (1 year, 5 months ago)

I was just curious,  but I don't blame you for not messing with it.


--------------------


Psilocybe cubensis data collection thread. please help with this project if you hunt wild cubensis.
https://www.shroomery.org/forums/showflat.php?Cat=0&Number=26513593&page=0&vc=1#26513593


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Offlinedelphin42
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Re: The Biopsy Method: A different way of doing LC. [Re: Josex]
    #26693035 - 05/24/20 06:22 PM (1 year, 4 months ago)

Thanks for this.  Just getting started.


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OfflineMH5109
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Re: The Biopsy Method: A different way of doing LC. [Re: delphin42]
    #26693129 - 05/24/20 07:04 PM (1 year, 4 months ago)

Iv been doing almost strictly biopsy transfers to new plates and it's a wonderful way to get a culture looking good. Im going to be doing a few LC runs here after I get the cultures looking good, its been a long time since I have used lc and only once has it turned out good.


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Invisiblebare.whiterabbit
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Re: The Biopsy Method: A different way of doing LC. [Re: MH5109]
    #26712537 - 06/01/20 11:01 PM (1 year, 4 months ago)

Krombipulos Mike forgot to bump this last time he saw it and couldn't remember what it was called, be he will be doing many rounds of this very soon. :super:

Quote:

Thrill said:
Bumping this cause it looks interesting and I got a question.

Anyone tried this through an injection port instead of shooting through a filter or opening and squirting it in? Or does pushing through the injection port rub off all the tissue cells?

Making a few jars of broth and thought Id give this a shot, see how it works vs some other methods that have popped up in the few years since I been away from here.




Shit, just saw this. He'll be doing his with SHIPs and a very small hole with double layered tyvek for minimal GE, so we'll let you know how it goes as soon as we can :thumbup:


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Edited by bare.whiterabbit (06/02/20 03:11 PM)


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Invisiblebare.whiterabbit
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Re: The Biopsy Method: A different way of doing LC. [Re: bongoman]
    #26712563 - 06/01/20 11:14 PM (1 year, 4 months ago)

Quote:

bongoman said:
Another question about this tek.

Josex says this is "a pour tek — don't aspirate the LC from the jar".

I wonder why not? What if I use a modded lid with an SHIP and a syringe filter, why can't I aspirate to a 60ml syringe with a 16G needle?




If you're doing an EzLC pull and then inoculating a larger volume, you can still flame your needle and hit up the larger jar. I think he pours, because he finds it reduces vectors for contam, but if you're more comfortable with SHIP use then by all means.

The idea behind re-flaming and then hitting a SHIP an additional time, is that it may not be totally clean itself and you run the risk of it contaminating your needle still.
But that's still the case regardless, if you're poking and transferring water into the original EzLC plate, so either way it's a toss up.

So long as you're cleaning everything properly and following principals you'll be fine doing a EzLC to SHIP transfer.

Krombipulos personally always pokes through one of those little packaged 70% alcohol swabs or you could use a small sponge soaked in iso. He's never had an issue with inoculation contams through a SHIP and does his in open air on his kitchen counter.


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Edited by bare.whiterabbit (06/02/20 03:12 PM)


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Invisibleretowen
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Re: Josex' Poke: A different way of doing LC. [Re: Josex]
    #26818368 - 07/12/20 12:11 AM (1 year, 3 months ago)

Definitely going to try this one - thank you for sharing!


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