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Offlinespandoos
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Advice needed - not getting anywhere
    #27067925 - 12/02/20 10:11 AM (3 years, 1 month ago)

Greetings,
I'm on my 5th attempt and thus far nothing has worked.

1st and 2nd attempts were with BRF cakes in 200ml jars. Mix was correct at 2:1:1 ratio. Pressure cooked at 15psi for 90min. inoculated with spore syringe inside a SAB. Temp was around 20-24C. Nothing happened for about 3 weeks. Then some white spots that stopped growing after 2 weeks. Shook the jars but nothing happed and in the end it grew mold.

Attempts 3,4 and 5
I got new syringes, made Agar which I left for a week to make sure it was still clean. inoculated the agar plates and once they were grown out I swithed to 500ml oats jars (boiled then simmered for 45min) air dried for 24hours then pressure cooked at 15psi for 90min.

It's been 3 weeks now and the oats jars started off well enough and once they got to about 30% I shook them. Again as previously the white growth spots are gone and what is their now is more gray with a gray milky discharge. Nothing looks what I have seen here or elsewhere online.

Any advice or ideas would be appreciated

https://files.shroomery.org/files/20-49/692879447-20201202_185247.jpg
https://files.shroomery.org/files/20-49/692879581-20201202_185259.jpg
https://files.shroomery.org/files/20-49/692879683-20201202_185313.jpg
https://files.shroomery.org/files/20-49/692879786-20201202_185345.jpg


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Offlinetiptrippy
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27067941 - 12/02/20 10:21 AM (3 years, 1 month ago)

If you have gotten clean cultures on agar then it sounds like the common issue is your sterilization process. What kind of PC are you using? Are you venting it properly?


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OfflineRoscoeReturnsS
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Re: Advice needed - not getting anywhere [Re: tiptrippy]
    #27067966 - 12/02/20 10:37 AM (3 years, 1 month ago)

Both your BRF jar and your grain jars look really wet to me. If too wet it can stall out your myc and promote contams.


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InvisibleIdiotCircusBoy
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Re: Advice needed - not getting anywhere [Re: tiptrippy]
    #27067970 - 12/02/20 10:40 AM (3 years, 1 month ago)

Don't shake BRF cakes.  The top verm layer acts as a barrier.  What do the tops of your lids look like?


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InvisiblebodhisattaMDiscordReddit
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Re: Advice needed - not getting anywhere [Re: IdiotCircusBoy]
    #27068002 - 12/02/20 11:07 AM (3 years, 1 month ago)

Those oat jars are prepared incorrectly. They're far too wet


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: tiptrippy]
    #27068044 - 12/02/20 11:39 AM (3 years, 1 month ago)

Quote:

tiptrippy said:
If you have gotten clean cultures on agar then it sounds like the common issue is your sterilization process. What kind of PC are you using? Are you venting it properly?




No idea what make it is but it's probably 30 years old. Doesn't have a weight on but a plug that extends upwards with marks denoting pressure.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: IdiotCircusBoy]
    #27068063 - 12/02/20 11:53 AM (3 years, 1 month ago)

Quote:

IdiotCircusBoy said:
Don't shake BRF cakes.  The top verm layer acts as a barrier.  What do the tops of your lids look like?




The BRF jars haven't been shaken at all and I wasn't planning to either. The lids have 4 holes in were the syringe went thru.

https://files.shroomery.org/files/20-49/693516727-20201202_205147.jpg

These 4 are 12 days old now.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: bodhisatta]
    #27068081 - 12/02/20 12:06 PM (3 years, 1 month ago)

Quote:

bodhisatta said:
Those oat jars are prepared incorrectly. They're far too wet




I was thinking the same thing but they were placed on the stove, brought up to boiling then back down to a simmer for 50min if I recall correctly. I left then for almost a full 24 hour spread out to cool off/dry before bottling them. At the time none clinged to my hands and their was a very few wet spots to nothing on the hand towels when i tested them.

I was thinking on throwing these out tomorrow and trying again to soak them for 24hours instead of using the stove...but I honestly don't know what I could be doing wrong.

Thinking on it now they look fine for about 2 weeks and it's usually around then perhaps 3 when I start noticing water building up on the bottom.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: RoscoeReturns]
    #27068100 - 12/02/20 12:18 PM (3 years, 1 month ago)

Quote:

RoscoeReturns said:
Both your BRF jar and your grain jars look really wet to me. If too wet it can stall out your myc and promote contams.




The BRF mix 2:1:1 i used the same coffee cup to measure everything so I can't see how I used to much water. Looking the the 4 jars bottoms I don't see the same water build-up as with the oats jars. I opened 1 jars lid and it smells kind of sour.


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Invisiblemushhead
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27068111 - 12/02/20 12:23 PM (3 years, 1 month ago)

a mere 100-200ml can throw off a brf jar easy.


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InvisibleIdiotCircusBoy
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27068222 - 12/02/20 01:27 PM (3 years, 1 month ago)

Your original post says you shook jar

Quote:

spandoos said:
1st and 2nd attempts were with BRF cakes in 200ml jars. Mix was correct at 2:1:1 ratio. Pressure cooked at 15psi for 90min. inoculated with spore syringe inside a SAB. Temp was around 20-24C. Nothing happened for about 3 weeks. Then some white spots that stopped growing after 2 weeks. Shook the jars but nothing happed and in the end it grew mold.





Additionally if those are your grain jar lids they need filter.  Where are you storing those jars during colonization?


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InvisibleJosex
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Re: Advice needed - not getting anywhere [Re: IdiotCircusBoy]
    #27068237 - 12/02/20 01:36 PM (3 years, 1 month ago)

Are those hulled oats?


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OfflineRoscoeReturnsS
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27068247 - 12/02/20 01:40 PM (3 years, 1 month ago)

Quote:

spandoos said:
Quote:

RoscoeReturns said:
Both your BRF jar and your grain jars look really wet to me. If too wet it can stall out your myc and promote contams.




The BRF mix 2:1:1 i used the same coffee cup to measure everything so I can't see how I used to much water. Looking the the 4 jars bottoms I don't see the same water build-up as with the oats jars. I opened 1 jars lid and it smells kind of sour.




You can have a perfect mix to start out, but depending on how deep the water is in your PC, and how you wrap them with foil, they can take on water during sterilization. I’ve had BRF jars come out saturated before.


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Invisiblemushhead
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Re: Advice needed - not getting anywhere [Re: Josex]
    #27068249 - 12/02/20 01:41 PM (3 years, 1 month ago)

What I see:
-jars too wet
-No visible holes for Gas Exchange.
Conclusion:
You need to properly vent your jars by adding punctures to the lids for Gas exchange.
EDIT:
I didn't see the fifth pic you posted showing punctured lids.
New Conclusion:
Your sterilization process is off, somehow there is moisture entering your jars during the PC cycle causing over hydration and subsequent contamination.
Possible point of contamination could be not using proper filters on your lids like Micropore tape or fiber disks.
One layer of Micropore tape over your holes will suffice if nothing else.


Edited by mushhead (12/02/20 01:49 PM)


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: IdiotCircusBoy]
    #27068363 - 12/02/20 02:31 PM (3 years, 1 month ago)

Quote:

IdiotCircusBoy said:
Your original post says you shook jar

Quote:

spandoos said:
1st and 2nd attempts were with BRF cakes in 200ml jars. Mix was correct at 2:1:1 ratio. Pressure cooked at 15psi for 90min. inoculated with spore syringe inside a SAB. Temp was around 20-24C. Nothing happened for about 3 weeks. Then some white spots that stopped growing after 2 weeks. Shook the jars but nothing happed and in the end it grew mold.





Additionally if those are your grain jar lids they need filter.  Where are you storing those jars during colonization?




The holes are covered by micropore tape. Everything is kept in a spare bedroom. Well lit but no direct sunlight. The temp is between 21C - 28C


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: Josex]
    #27068387 - 12/02/20 02:40 PM (3 years, 1 month ago)

Quote:

Josex said:
Are those hulled oats?




Should be?
https://files.shroomery.org/files/20-49/694519151-20201202_233438.jpg


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InvisibleJosex
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27068394 - 12/02/20 02:42 PM (3 years, 1 month ago)

No, you want the hulls intact like in the pic. They look so wet and mushy in the OP that I thought for a moment they could be hulled oats.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: RoscoeReturns]
    #27068398 - 12/02/20 02:48 PM (3 years, 1 month ago)

Quote:

RoscoeReturns said:
Quote:

spandoos said:
Quote:

RoscoeReturns said:
Both your BRF jar and your grain jars look really wet to me. If too wet it can stall out your myc and promote contams.




The BRF mix 2:1:1 i used the same coffee cup to measure everything so I can't see how I used to much water. Looking the the 4 jars bottoms I don't see the same water build-up as with the oats jars. I opened 1 jars lid and it smells kind of sour.




You can have a perfect mix to start out, but depending on how deep the water is in your PC, and how you wrap them with foil, they can take on water during sterilization. I’ve had BRF jars come out saturated before.




I fill the PC with about an inch of water. If more it doesn't come past the bottom 1cm of the jars. All holes are micropore taped and then covered with foil. I don't recall ever noting the micropore tape being wet when removing the foil or seeing water on the lid under the foil either but I'll take special note on the next round


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Invisiblemushhead
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27068403 - 12/02/20 02:51 PM (3 years, 1 month ago)

Ah so you're adding your micropore tape on and then PCing the jars.
There I believe is your issue. You need to PC without micropore and add it to the jars at the time of inoculation.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: mushhead]
    #27069121 - 12/02/20 11:02 PM (3 years, 1 month ago)

Quote:

mushhead said:
Ah so you're adding your micropore tape on and then PCing the jars.
There I believe is your issue. You need to PC without micropore and add it to the jars at the time of inoculation.




Ok I'll try that on the next batch. I use micropore tape since I couldn't find polyfill easily. But from what I've seen lids with polyfill are PC'd with them in so how can micropore closed holes make such a big difference?

I used my last bit of Agar to make some LC since Im still waiting on another batch of sterilised petri dishes.
The syringes I received are 10ml in size with 8ml LC.
How much LC per hole on the oat jars?
1ml/2ml
For the 200ml BRF jars with 4 holes?
1ml/2ml
I recall using 1ml per hole since I used a 8ml for 2 jars


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Invisiblebigfootscreepyuncl
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Re: Advice needed - not getting anywhere [Re: mushhead]
    #27069136 - 12/02/20 11:30 PM (3 years, 1 month ago)

Quote:

mushhead said:
Ah so you're adding your micropore tape on and then PCing the jars.
There I believe is your issue. You need to PC without micropore and add it to the jars at the time of inoculation.




I'm not trying to hijack a thread, so I apologize in advance. I'll be doing my first BRF tek tomorrow and have been trying to find the answer to this and keep finding differing answers.

I'll be 'steaming' the cakes, not in a PC since Santa wont let me open my PC until Christmas morning...Anyway, what you're saying is that I should fill the jars, cover with foil, steam, move to SAB, inoculate THEN place the tape over the holes? Or tape the holes before I inoculate? This option seems like there's more risk of contamination than doing the tape after inoculation.

Sorry for asking such a dumb question but I've seen it every which way and I'd like this attempt to be successful. Cheers, mate!


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OfflineRoscoeReturnsS
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Re: Advice needed - not getting anywhere [Re: bigfootscreepyuncl]
    #27069245 - 12/03/20 02:46 AM (3 years, 1 month ago)

Quote:

bigfootscreepyuncl said:
Quote:

mushhead said:
Ah so you're adding your micropore tape on and then PCing the jars.
There I believe is your issue. You need to PC without micropore and add it to the jars at the time of inoculation.




I'm not trying to hijack a thread, so I apologize in advance. I'll be doing my first BRF tek tomorrow and have been trying to find the answer to this and keep finding differing answers.

I'll be 'steaming' the cakes, not in a PC since Santa wont let me open my PC until Christmas morning...Anyway, what you're saying is that I should fill the jars, cover with foil, steam, move to SAB, inoculate THEN place the tape over the holes? Or tape the holes before I inoculate? This option seems like there's more risk of contamination than doing the tape after inoculation.

Sorry for asking such a dumb question but I've seen it every which way and I'd like this attempt to be successful. Cheers, mate!




Are you doing the standard PF tek with a dry verm layer at the top of the jar? If so, don’t bother with the tape. It’s not necessary. Using MP tape as your only filter is not really the best way to go about this. It works well on tubs to regulate airflow, but a single layer on jars has been known to fail and allow contaminants in. Some use multiple layers, but that stuff gets kinda gross in the PC. If you use the dry verm layer, that is your filter and you don’t need anything else. If you don’t use the dry verm layer, or you’re doing grain jars, I would suggest SFDs or poly fill over MP tape.


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OfflineOvoidhunter
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Re: Advice needed - not getting anywhere [Re: RoscoeReturns]
    #27069260 - 12/03/20 03:27 AM (3 years, 1 month ago)

Your culture wasn't clean to begin with if you didn't do any transfers from the germination dish. Going straight from the germ dish is the same as not cleaning at all and injecting directly from the syringe.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: Ovoidhunter]
    #27070213 - 12/03/20 03:12 PM (3 years, 1 month ago)

Quote:

Ovoidhunter said:
Your culture wasn't clean to begin with if you didn't do any transfers from the germination dish. Going straight from the germ dish is the same as not cleaning at all and injecting directly from the syringe.




I've used a bought syringe to BRF. Then syringe to agar to oats and used my last agar plate to create LC and used that on my last run. On the 40 plates I had 5 with contaminants but the rest all looked good.
The LC is clear with only the cloudy mys at the bottom.

Since my PC is very old...I'm wondering now if it is in fact reaching 15psi and if not is that's perhaps causing the extra water buildup.

Im ready for another run but I just can't see how low PSI could cause the extra wetness nor after 5 attempts that I've been unable to properly use foil :confused:


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OfflineFrugalFungi
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Re: Advice needed - not getting anywhere [Re: spandoos]
    #27070610 - 12/03/20 07:20 PM (3 years, 1 month ago)

Those oats are way to wet my friend.


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Offlinespandoos
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Re: Advice needed - not getting anywhere [Re: FrugalFungi]
    #27083261 - 12/10/20 11:58 PM (3 years, 1 month ago)

I've made up a new batch of oats. Boiled cooked them for 50min. Left to air dry for 24hours theb placed in the fridge as per BOBs oats prep tek for 12 hours then PC'ed for 90min.

I can see some condensation inside and some oats sticking to the sides but most move around freely as I turn the jar.

How does this look to the experienced eye since I can't see any difference to the previous attempts.

https://files.shroomery.org/files/20-50/766960317-20201211_084639.jpg
https://files.shroomery.org/files/20-50/766960450-20201211_084625.jpg
https://files.shroomery.org/files/20-50/766960592-20201211_084615.jpg
I want to inoculate with LC - thinking of 1CC per jar? These are 500ml jars.

Thanks.


Edited by spandoos (12/10/20 11:59 PM)


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