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OfflineAdustus
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Starting my mushrooms in some agar
    #19253302 - 12/09/13 11:46 PM (10 years, 1 month ago)

So im going to use PastyWhytes Agar tek and use some live cultures I have.

I have a PC and made a SAB and have all the sterilizing stuff.

Anyone have any advice for a first timer using agar? And how to maybe make it go smoother? Im using jars instead of gladware.

I also was wondering if anyone had any ideas for a rack? My lid dont seem to wanna fit inside very well.

These are my only cultures so im really concerned with the inoculation process bringing contams. Any help is greatly appreciated!


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OfflineForresterM
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19254542 - 12/10/13 09:45 AM (10 years, 1 month ago)

The biggest help I've found is once you have your SAB wiped down with alcohol and everything all set up in there and ready to go, leave it to sit for like an hour so any spores can have a chance to settle.  I used to do 15 minutes because that's what I read, but I had a lot of contamination problems until I just started waiting longer.  Also try to minimize the number of times you bring your hands in and out of the box. 

If you're using ball jars like me, get your lid rings broken loose before you let it sit, because it's tough to break them loose in the box with your hands covered in alcohol and slippery.  You wanna have everything placed right and a plan in your head of exactly what you're gonna do, do it quick and minimize the time you have the jars open.

Most people around here flame their scalpel blade between cultures, but I just use a new one so I'm not bringing my hands in and out of the box with hot blades and creating air currents.  They're like $7 for a box of 100.

I'm no agar expert these are just a few things I've done to get much better results in my SAB.  Also, I use the upside down box on top of moist towel routine that Notahacker420 shows here, this way any spores that fall stick to the towel and don't get stirred up again.


--------------------
Repugnant is a creature who would squander the ability to lift an eye to heaven, conscious of his fleeting time here.
-------------------

Have some medicinal mushrooms and want to get the most out of them?  Try this double extraction method.


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OfflineAmanita virosa
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Re: Starting my mushrooms in some agar [Re: Forrester]
    #19255130 - 12/10/13 12:17 PM (10 years, 1 month ago)

Although jars work fine, consider investing 7 bucks in a sleeve of sterile plastic petri plates.  MUCH easier to work with, esp. in an sab.  take up less room in the fridge, less awkward to sub out of and easier to see contam.  Otherwise, allowing your jars to cool with the lids slightly loose (flip the lids upside down, rubber side up) will allow excess moisture to escape.  Otherwise, you may have some on the surface of the agar.  Consider making some slants for long term storage of your cultures.


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OfflineAdustus
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Re: Starting my mushrooms in some agar [Re: Forrester]
    #19256266 - 12/10/13 04:37 PM (10 years, 1 month ago)

Quote:

Forrester said:
The biggest help I've found is once you have your SAB wiped down with alcohol and everything all set up in there and ready to go, leave it to sit for like an hour so any spores can have a chance to settle.  I used to do 15 minutes because that's what I read, but I had a lot of contamination problems until I just started waiting longer.  Also try to minimize the number of times you bring your hands in and out of the box. 

If you're using ball jars like me, get your lid rings broken loose before you let it sit, because it's tough to break them loose in the box with your hands covered in alcohol and slippery.  You wanna have everything placed right and a plan in your head of exactly what you're gonna do, do it quick and minimize the time you have the jars open.

Most people around here flame their scalpel blade between cultures, but I just use a new one so I'm not bringing my hands in and out of the box with hot blades and creating air currents.  They're like $7 for a box of 100.

I'm no agar expert these are just a few things I've done to get much better results in my SAB.  Also, I use the upside down box on top of moist towel routine that Notahacker420 shows here, this way any spores that fall stick to the towel and don't get stirred up again.




Good advice Ill keep the exposure to a minimum:thumbup:


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OfflineAdustus
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Re: Starting my mushrooms in some agar [Re: Amanita virosa]
    #19256270 - 12/10/13 04:38 PM (10 years, 1 month ago)

Quote:

Amanita virosa said:
Although jars work fine, consider investing 7 bucks in a sleeve of sterile plastic petri plates.  MUCH easier to work with, esp. in an sab.  take up less room in the fridge, less awkward to sub out of and easier to see contam.  Otherwise, allowing your jars to cool with the lids slightly loose (flip the lids upside down, rubber side up) will allow excess moisture to escape.  Otherwise, you may have some on the surface of the agar.  Consider making some slants for long term storage of your cultures.





Petris are my next investment. If you have a good source to buy a sleeve id appreciate it my friend.


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OfflineShu
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19256350 - 12/10/13 04:55 PM (10 years, 1 month ago)

Move very slowly in the SAB, air currents are the enemy.

Open your plates only far enough to pass in your culture and close them quickly.

Practice your transfers using empty containers and no cultures to build up muscle memory.

Transferring smaller pieces of culture to many separate dishes increases your odds of a clean transfer.

Have a few spare plates ready for second pass in case all your plates contaminate and pass the culture at the first sign of contam.

Never open a moldy plate in your SAB.

What "lid" are you speaking of?


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OfflineAdustus
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Re: Starting my mushrooms in some agar [Re: Shu]
    #19264172 - 12/12/13 03:08 AM (10 years, 1 month ago)

Quote:

Shu said:
Move very slowly in the SAB, air currents are the enemy.

Open your plates only far enough to pass in your culture and close them quickly.

Practice your transfers using empty containers and no cultures to build up muscle memory.

Transferring smaller pieces of culture to many separate dishes increases your odds of a clean transfer.

Have a few spare plates ready for second pass in case all your plates contaminate and pass the culture at the first sign of contam.

Never open a moldy plate in your SAB.

What "lid" are you speaking of?




Meant lids from my jars.


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OfflineSkysTheLimit
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Re: Starting my mushrooms in some agar [Re: Shu]
    #19264225 - 12/12/13 03:55 AM (10 years, 1 month ago)

Quote:

Shu said:
Move very slowly in the SAB, air currents are the enemy.

Open your plates only far enough to pass in your culture and close them quickly.

Practice your transfers using empty containers and no cultures to build up muscle memory.

Transferring smaller pieces of culture to many separate dishes increases your odds of a clean transfer.

Have a few spare plates ready for second pass in case all your plates contaminate and pass the culture at the first sign of contam.

Never open a moldy plate in your SAB.

What "lid" are you speaking of?




I think you should try spraying down the glovebox with 10% bleach or lysol or w/e after you put all your work inside. The water vapour traps dust and spores causing them to stick to surfaces.
After an hour or so, you can assume most of the dust/spores in your SAB has either settled down or stuck to a surface.

I do this and I'm pretty sure most SAB users do the same, its also done in plant cloning where the specimens are just as vulnerable to contamination in mycology.


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Offlinebjanuszewski
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Re: Starting my mushrooms in some agar [Re: SkysTheLimit]
    #19265141 - 12/12/13 10:24 AM (10 years, 1 month ago)

if your worried about contam you need to make sure you have clean tissue thats been handle very little. you can always isolate away from contam too.

my greatest success with agar comes from using 1/2 pint jars w/ tyvek over AE hole and using my colonized rye berries to inoc the agar. using tissue can be hard bc you'll usually have to isolate to get nice clean looking mycelium from my experience.


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OfflineAdustus
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19270911 - 12/13/13 02:01 PM (10 years, 1 month ago)

Well it seems to have gone well so far. I spray like all hell through every step with 91% alcohol or peroxide. My gas exchange tape melts to my jars in the PC and I think my rack got all fucked up too.

Do i need more water in my PC? I fill it to just cover the jar lids.

Anyone have any ideas for gas exchange? I tap two holes with nails in the lid and planned to use the tape but it doesnt seem to like the PC. Will leaving the holes open allow too much moisture inside?

I just need to figure out my gas exchange now :/


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Invisibleblindingleaf
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19271227 - 12/13/13 03:16 PM (10 years, 1 month ago)

if u r using 1/2 pint glass jars u can just turn the lid disc upside down (even that is not needed though), agar barely need GE (though it may be debated here, I have found it does not)  I think you should search "Doc-T's no pour Tek" because he uses 1/2 pint glass jars and if I remember correctly, no MOD for GE, just put the law disc and ring on normal with no holes.  if u already have holes in the lid, go with Tyvec like bj said, but don't cook them with exposed holes, WAAAY too much moisture will get in...  u don't need more water in PC, less than if u are cooking grains because the PC will only run 30-35 minutes.  also, don't put too much agar in each jar, because its annoying to transfer something thicker than 1/4 inch, I'd say go for 1/8 inch, u don't need much, just a light covering of the bottom.  In my AA if I put water to JUST under the rack disc it came with, thats more than enough to run 30-35 minutes.  Good Luck!  Agar is fun.  I think Im better with agar than actually growing shrooms!  its addicting, get some PP5s and ill have a full time hobby, as if growing wasn't enough right?


--------------------
A few thoughts on cultivation
MICROBIAL HUSBANDRY!!!!

The whole is greater than the sum of its parts


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Invisibleblindingleaf
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Re: Starting my mushrooms in some agar [Re: blindingleaf]
    #19271235 - 12/13/13 03:17 PM (10 years, 1 month ago)

also, my micropore tape is fine in PC, esp if just for 30 min, is that the tape u were referring to that did not do well in PC?


--------------------
A few thoughts on cultivation
MICROBIAL HUSBANDRY!!!!

The whole is greater than the sum of its parts


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OfflineAdustus
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Re: Starting my mushrooms in some agar [Re: blindingleaf]
    #19273560 - 12/14/13 06:24 AM (10 years, 1 month ago)

Quote:

blindingleaf said:
also, my micropore tape is fine in PC, esp if just for 30 min, is that the tape u were referring to that did not do well in PC?




Yeah I got some weird cheap shit I found online. Well Im new to agar and I made way too much from what it sounds. I bet my reishi culture I traded for is fucked. Ill have to review that tek and take another shot at agar. Do you use food coloring in yours to spot contams better?


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OfflineForresterM
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19273628 - 12/14/13 07:09 AM (10 years, 1 month ago)

If the tape isn't working out, you could always use post-office tyvek.  Holds up well in the PC.  Just silicon a little patch over whatever hole you've got in the lid.


--------------------
Repugnant is a creature who would squander the ability to lift an eye to heaven, conscious of his fleeting time here.
-------------------

Have some medicinal mushrooms and want to get the most out of them?  Try this double extraction method.


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OfflineAdustus
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19273683 - 12/14/13 07:47 AM (10 years, 1 month ago)

Badass! I just found out about the pillow stuffing as a means of gas exchange. I have a couple tubes thatll do fine.


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OfflineForresterM
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19273895 - 12/14/13 09:37 AM (10 years, 1 month ago)

Yeah polyfill works too.  Just cover the lids with foil in the PC.


--------------------
Repugnant is a creature who would squander the ability to lift an eye to heaven, conscious of his fleeting time here.
-------------------

Have some medicinal mushrooms and want to get the most out of them?  Try this double extraction method.


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OfflineAmanita virosa
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Re: Starting my mushrooms in some agar [Re: Adustus]
    #19273903 - 12/14/13 09:41 AM (10 years, 1 month ago)

Quote:

Adustus said:
Well it seems to have gone well so far. I spray like all hell through every step with 91% alcohol or peroxide. My gas exchange tape melts to my jars in the PC and I think my rack got all fucked up too.

Do i need more water in my PC? I fill it to just cover the jar lids.

Anyone have any ideas for gas exchange? I tap two holes with nails in the lid and planned to use the tape but it doesnt seem to like the PC. Will leaving the holes open allow too much moisture inside?

I just need to figure out my gas exchange now :/




Too much water!  Two inches of water is all you need in the pc. Just be sure to vent the air out till pure steam is coming out before putting the weight on. If you put holes in the lid and the tape melted they are toast. Micro pore tape makes a terrible filter. Honestly, you could leave out the gas exchange altogether. Just use plastic lids. They are not air tight. Or even better buy some dang petri plates.  You can re sterilize the agar. It will liquefy when heated.


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Offlinet3chnobily
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Re: Starting my mushrooms in some agar [Re: Forrester]
    #19273910 - 12/14/13 09:42 AM (10 years, 1 month ago)

pack the polyfill tight if you want it to be an effective filter.  I use tyvek squares taped down with 3m micropore and they last many pc cycles.


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Invisibleblindingleaf
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Re: Starting my mushrooms in some agar [Re: t3chnobily]
    #19275145 - 12/14/13 03:11 PM (10 years, 1 month ago)

amanita...I have been curious to try not putting a ge holes in these containers..



I was under the impression that without one they would pop in the PC...do u think the pressure inside would just slowly push the air out instead, even though its a snap on?  for the screw on PP5 I just loosen them a bit, no GE hole.


--------------------
A few thoughts on cultivation
MICROBIAL HUSBANDRY!!!!

The whole is greater than the sum of its parts


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OfflineAmanita virosa
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Re: Starting my mushrooms in some agar [Re: blindingleaf]
    #19275217 - 12/14/13 03:30 PM (10 years, 1 month ago)

Dunno. Have you tried nuking them?  If the are not polypropylene or some comparable plastic they will melt. I suspect the lids would  pop right off.


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