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Offlineavaratz
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Registered: 10/18/20
Posts: 7
Last seen: 2 years, 11 months
Getting started with agar
    #26999121 - 10/23/20 05:25 AM (3 years, 3 months ago)

Hi all,

After an absolute disaster of my attempt at BRF, I've ordered what I need for agar and I'm going to give it a go.

Would a more experienced, kind person please take a look through my checklist to see if I'm missing anything/understanding anything incorrectly? It would really set my mind at ease and I'd really appreciate it! :smile:

1. Follow https://www.shroomery.org/forums/showflat.php/Number/25137693/fpart/all/vc/1 to make no-pour agar plates
    a. Small holes in side of plastic containers
    b. 3x layers of micropore over hole
    c. 2% agar, 2% light malt extract, green food colouring in agar    mix
    d. Pour a thin layer into each container
    e. Pressure cook containers, remove from PC when safe and set on counter for level set of agar
    f. Wait at least 3 hours, until fully cool and set
2. Prepare my SAB
    a. 70% iso wipe down of work surface and sides/roof of SAB
    b. Spray inside of SAB with soapy water and allow to settle
    c. Place agar plates inside SAB, left hand side of container, upside down
    d. Place something (after sanitising it) to elevate plates above the work surface
3. Shower, brush teeth, fresh clothes
4. Gloves and mask – wipe down gloves with 70% iso every time something is touched outside the SAB
5. Wipe down outer packaging of spore syringe, remove. Wipe down syringe and shake well
6. Inside the SAB, attach needle to syringe
7. Flame the syringe using my gas hob until the needle is red hot (outside the SAB)
8. Spray out a little of the syringe until sizzling stops to cool the needle
9. Back inside the SAB now, pick up a plate and place it on my elevated surface
10. Open the lid of the plate while upside down and as quickly as possible turn the plate around and place a single drop of spore syringe in the middle of the plate
11. Replace the lid and place the plate (right side up this time so the spores can settle) to the right-hand side of the container, forming a new stack on the opposite side of the uninoculated stack.
12. Repeat steps 7 to 11 for remaining plates
13. Store plates somewhere between 18 – 24 C (64 – 75 F)


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OfflineRoscoeReturnsS
Crotchety chode man
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Registered: 02/12/18
Posts: 1,756
Loc: State of Confusion
Last seen: 5 hours, 11 minutes
Re: Getting started with agar [Re: avaratz]
    #26999146 - 10/23/20 05:57 AM (3 years, 3 months ago)

Looks good. One change, on 7, 8 & 9. After flaming the needle, bring it in the SAB before cooling it. You don’t want a cool needle in open air.


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OfflineCaptainParanoid
Stranger Danger
I'm a teapot User Gallery


Registered: 08/06/20
Posts: 74
Last seen: 1 year, 5 months
Re: Getting started with agar [Re: RoscoeReturns]
    #26999168 - 10/23/20 06:20 AM (3 years, 3 months ago)

I don’t have lots of agar experience but I’ve been pouring my own plates, inoculating, and doing transfers with a good success rate.

I like to use tyvek arm gauntlets (spray and wipe with alcohol like gloves) and sterilized rubber gloves (one set) individually wrapped. But I can get all that I need for free at work. I’m sure the gloves aren’t cheap but I have no idea.

I’m surprised more people don’t report using the tyvek gauntlets though.


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Offlineavaratz
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Registered: 10/18/20
Posts: 7
Last seen: 2 years, 11 months
Re: Getting started with agar [Re: RoscoeReturns]
    #26999211 - 10/23/20 07:05 AM (3 years, 3 months ago)

Thanks, good point! Could be why my BRF cakes ended up contammed...


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Offlineavaratz
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Registered: 10/18/20
Posts: 7
Last seen: 2 years, 11 months
Re: Getting started with agar [Re: CaptainParanoid]
    #26999214 - 10/23/20 07:07 AM (3 years, 3 months ago)

Quote:

CaptainParanoid said:
I don’t have lots of agar experience but I’ve been pouring my own plates, inoculating, and doing transfers with a good success rate.

I like to use tyvek arm gauntlets (spray and wipe with alcohol like gloves) and sterilized rubber gloves (one set) individually wrapped. But I can get all that I need for free at work. I’m sure the gloves aren’t cheap but I have no idea.

I’m surprised more people don’t report using the tyvek gauntlets though.




Something like these, combined with latex gloves?

https://www.ebay.co.uk/p/1272975736


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OfflineRoscoeReturnsS
Crotchety chode man
Male


Registered: 02/12/18
Posts: 1,756
Loc: State of Confusion
Last seen: 5 hours, 11 minutes
Re: Getting started with agar [Re: CaptainParanoid]
    #26999237 - 10/23/20 07:36 AM (3 years, 3 months ago)

Quote:

CaptainParanoid said:

I’m surprised more people don’t report using the tyvek gauntlets though.




Probably because they really are not necessary. I used them when I started out. Then I stopped, and still had really good results. Sterile technique is so much more important. Now I rarely even use gloves. Wash well before starting, ISO wipe down, keep my hands and arms from going above an open plate or jar and no problems. Plus, unless you wash, clean, or replace your tyvek sleeves, they end up being full of contams and dead skin just waiting to fall off in your SAB.


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Invisiblecoversall
إِنْ شَاءَ ٱللَهُ
 Unread Journal


Registered: 06/06/20
Posts: 2,749
Loc: संसार
Re: Getting started with agar [Re: RoscoeReturns]
    #26999281 - 10/23/20 08:13 AM (3 years, 3 months ago)

Same, my hands and forearms as usually wet with ISO when I'm working in my SAB. I get a lot of nicks on my hands and forearms at work, so using ISO like this means I haven't had an infected cut in ages!


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

..:: E V E R Y  ::..

..:: New? Start here. ::..
..:: How I Panaeolus. From Agar to Tea ::..


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OfflineCaptainParanoid
Stranger Danger
I'm a teapot User Gallery


Registered: 08/06/20
Posts: 74
Last seen: 1 year, 5 months
Re: Getting started with agar [Re: avaratz]
    #26999419 - 10/23/20 10:11 AM (3 years, 3 months ago)

Quote:

avaratz said:

Something like these, combined with latex gloves?

https://www.ebay.co.uk/p/1272975736





Yes, that’s what I use. I also don’t worry about showering and changing my clothes. At the end of the day there is no being too sterile. Are gauntlets necessary...definitely not. I try to keep anything going into my SAB as sanitized as possible. Tyvek is easier to keep clean over my bare hairy arms and more forgiving if I make a mistake such as putting my arm over an open plate or jar or whatever.
As I said though. I get all the Tyvek gauntlets, steril gloves, hair nets and face coverings I want for free. I could even get sterilized clean room jump suits. I only wish I could use the clean room labs for my hobby.


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OfflineShakedown Street
Friend
Male

Registered: 09/05/20
Posts: 383
Last seen: 3 months, 9 days
Re: Getting started with agar [Re: CaptainParanoid]
    #27001520 - 10/24/20 04:20 PM (3 years, 3 months ago)

A glass media bottle like the 500 ML GL45 round media bottle
to pasteurize your agar in the PC.  You could use a 750 mL glass bottle, but you have to cap it with a wad of polyester fluff & make sure it doesn't fall over in the PC.  This glass media bottle is shorter
& squatter and makes the job easier even though it only holds 500 mL instead of 750.  But that is enough to do many agar dishes.

You don't need micropore-covered holes in the sides of the containers of agar.  They don't need to breathe.


Edited by Shakedown Street (10/24/20 04:31 PM)


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