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InvisibleD3_Myc
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D3's Perfect transfers and agar TEK. * 65
    #26628234 - 04/25/20 09:35 PM (3 years, 8 months ago)

Perfect Transfers and Agar TEK


I'm frequently asked how I get my agar so clear, the colors and how I get such perfect transfers. The following below is a detailed explanation of the process. There's many ways to achieve similar results but this what works for me. Feel free to experiment with different recipes and nutrient levels.



Tools Required






Petri Dishes




By far Polystyrene and Glass will give the best clarity. I use polypropylene for streaks or other circumstances where I don't expect to show off samples. Nothing beats the cheap disposable dishes though. I've tried no pours, deli cups, glad mini rounds.. none where for me. The particular brand I'm using right now is Celltreat. I'm open to others suggestions on brands. Right now a case is averaging me around $100 for 500 count.

The Agar


I've tried several brands of agar agar and premade mixes. Most left fibrous bits in my petris detracting from the overall appearance. Ultimately I have found that this brand gives consistent results



For nutrients i'm using Maltoferm light malt extract it disolves clean and clear.



For coloring, Misty turned me onto this brand. Wilton Neon Gel it has given best results and color doesn't break down in the PC. But after continued testing and anecdotal evidence it has been found that the red and purple dyes contain red#3 or erythrosin which is naturally fungicidal. It’s caused abnormal, stunted or sporadic growth and my recommendation to avoid those two colors or any other dyes that contain red#3 I’ll be doing a side by side comparison and updating a link here for future users. (Updated 10-22-20) Here's the current link on the experiment thread



Another great brand is Chef Master Neon Gel


Another option is pouring black plates using this Charcoal Powder approximately 1g per 100ml but honestly I don't even measure.

9-15-21 New link for charcoal



Ingredients


The following recipe will produce about 40 petri dishes. You can reduce by half if pouring only one sleeve of 20. I've had best results so far with this amount of nutrients, feel free to experiment with more or less levels of LME. Lower the nutrients the more likely you will get rhizomorphic growth, higher nutes more tomentos. Of course genetics and other environmental conditions come into play as well.

  • 20g agar agar
  • 16g LME
  • 1000ml distilled water


Edit 8-4-21

To switch things up can also add 1g nutritional yeast to the above recipe. I like to pour plates of each and send transfers to both medias.

Note: when using nutritional yeast, agar will become cloudy. I recommend using the charcoal powder for yeast plates.

Directions


1. Add distilled water to pot and begin heating to light boil.
2. While water is heating up, measure out dry ingredients.
3. Once water is beginning to lightly boil, whisk in dry ingredients and reduce heat to prevent boil over.
4. If you have a stick blender, blend for 30-45 seconds thoroughly.



5. Continue to lightly boil agar for 1-2 minutes stirring to prevent foaming and boil over.
6. Add food coloring to media bottles. Personally I use 3 bottles for a 1000ml batch. 3 drops food coloring or 1 drop per 100ml per media bottle. Have fun, experiment with mixing colors.
7. Funnel agar mix into media bottles distributing evenly among bottles. Swirl to distribute food coloring, cap loosely and cover top with foil if you desire. After a few years I’ve pretty much eliminated using foil for most things.



Edit 8-4-21

8. Add enough water to PC for a 20 minute run. For fairly cheap you can get a raised rack for your PC keeps your jars and media bottles out the water. Fill PC to just under or touching the rack.



9. Follow the manufacturer instruction for purging and bringing PC up to a boil prior to adding the weight. I use the modified weight with 2 quarters tapped to it. This reduces rattle and allows the pressure cooker to reach pressures above 15psi.
10. Bring the PC up to 15-17psi and set timer for 20 minutes. Continue to monitor pressure don't allow it to exceed 17psi for safety sake.
11. Once time is up, turn off heat and allow pressure to release naturally, use this time to shower, prep SAB if using one, wipe down your prep surfaces ect.
12. In pot large enough to hold media bottles, heat several inches of water to 140f (60c) or if you own a sous vide this is even better.
13. Carefully remove media bottles from PC, place into pot of water ( this serves two purposes, one it helps cool the bottles down to handling temps and helps keep bottles warm to prevent from setting up while pouring other bottles) move your pot and media bottles to where you will be performing pours.
14. Using a laser temp probe measure the media bottles temperature. I begin pouring once temperature reaches 135-140f (57-60c)

Once all plates are poured leave unwrapped in SAB with arm holes covered for several days or several hours in front of flowhood to allow condensation to subside.

 

The end result is a crystal clear plate with no condensation




Taking Transfers


This method of taking transfers can be performed within a SAB or in front of a flow hood. For obvious reasons a flow hood is much easier to work in front of.



Tools Needed


The main focus of this method utilizes a punch or stainless steel straw to act as a cookie cutter making perfectly round holes in your agar for transfer. The method has been floating around on the groups for some time now. You can make one these out of brass tubes, metal straws what ever. You're only limited by your imagination. I’ve been using a custom punch made with brass tubing and has an acrylic handle.



  • Leather hole punch
  • Flowhood or SAB
  • Scalpel
  • Torch or alcohol burner
  • Parafilm or cling wrap... what ever you prefer wrapping your plates with
  • Petri dishes transferring from.
  • Poured petri dishes for transferring to.
  • Spare dish for cooling heated tools ( I use a ugly dish, one that had sediment or perhaps agar started to gel toward end of pours)



Directions


1. Select the plates you will be transferring from.
2. Prep you work surfaces as usual, wipe down with iso ect.
3. Flame sterilize your scalpel and place into a container blade up.
4. Flame sterilize your punch.You will not be able to get the steel red hot without burning yourself and not being able to handle the tool. Just gently flame the tip and slot opening for a few good seconds like shown in video above.
5. Cool the leather punch / transfer tool in the spare petri dish by gently depressing into agar several times until it stops sizzling. If using brass tubing, ss straw ect this will be not necessary as the metal cools fairly quickly on its own.
6. Use the punch to cut perfect lil circles into the leading edge of your myc





7. Using your previously flame sterilized scalpel, gently poke into the side of your transfer disc and lift. This may take a little practice but it's extremely easy once you have done it a couple times. i recommend practicing on a blank petri until you get the hang of it.







By gently pressing the disc onto the recieving plate it should stick and allow you to slide your scalpel out. Sometimes if your scalpel slipped too far into the disc you may have a little difficulty getting it free.

You're left with a perfect little transfer disc in the center of your agar.



The end result after a few transfers are beautifully symmetrical dishes.





Hope this little tutorial helps you elevate your agar and transfer game. I'll update periodically with better videos and pictures.

Thanks, D3


Edited by D3_Myc (10/22/21 08:03 PM)


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InvisibleD3_Myc
Weeb Trash
I'm a teapot


Registered: 05/06/18
Posts: 4,399
Loc: Year Zero
Trusted Cultivator
Re: D3's Perfect transfers and agar TEK. [Re: pureshrooming]
    #26628326 - 04/25/20 10:19 PM (3 years, 8 months ago)

I need to fix some the picture orientations. Looked better on the laptop


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InvisibleD3_Myc
Weeb Trash
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Registered: 05/06/18
Posts: 4,399
Loc: Year Zero
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Re: D3's Perfect transfers and agar TEK. [Re: hallmaker]
    #26628454 - 04/25/20 11:36 PM (3 years, 8 months ago)

Quote:

hallmaker said:
Nice right up duuuuude. Gonna definitely use the food coloring and leather punch tips. Couple quick questions.
1. In a earlier post, I seen you cross breed using liquid streaks. Would putting multiple agar punches from 2 different strains do the same? Also, I'm gonna use the marker on the petri dish lid to trace the streak tip.
2. Do you do anything special with your camera for clearer shots. Both my camera and phone blur just before the shot
3. Is your leather punch particularly extra sharp or will a decent set from Amazon be ok?

Thanks for the help




Any set should be fine, there’s also other options like dermal punches. I got a tool being made for me right now that has a custom made handle of either wood or acrylic. Look forward to trying it out.

I’ve never used liquid inoculant for streaks only spores. I traced my streaks out and labeled them before hand to just help
Me plan and remember what was streaked where since I was doing so many different cross attempts.



As far as the sandwiching agar pucks for crosses I’m not sure. Probably a better question for the more experienced folks over on the strain crossing thread.


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InvisibleD3_Myc
Weeb Trash
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Registered: 05/06/18
Posts: 4,399
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Re: D3's Perfect transfers and agar TEK. [Re: orange771]
    #26628459 - 04/25/20 11:41 PM (3 years, 8 months ago)

Quote:

orange771 said:
Ive been using a cork borer, cheap red one off of amazon about $7 and it works great. Basically does what the leather punch and scalpel do but with one tool.




I just looked up the cork borer, interesting tool. Does the puck stick inside it when you do the punch then you just pop it out with the t-handle? If so curious as to how it stays inside. I think I’ve only had it stick like once with the leather punch. Would like to see it in action.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: gizmo1]
    #26628537 - 04/26/20 12:25 AM (3 years, 8 months ago)

Quote:

gizmo1 said:
That's some really good looking growth 👌
D you havent had any problems with contams dropping of your hand onto the open plate or the piece being transferred?
I get contams everytime I accidentally pass my gloved hand over a plate. Especially when I'm pouring. I always know which sleeve is going to have a bad plate in it. I see the pic where you are cutting the disk and it scares the shit outta me lmao. I mean it seems to be working good for you but it makes me uncomfortable for some reason.




The only contams I’ve gotten where from spore streaked plates. I don’t even wrap the cooling plate and have never had it contam. I’ll leave the plate out for a few weeks or until it’s hideously scarred from me cooling tools in it. Not sure if that’s a testament to how clean my lab is or what. I do frequently squirt some iso on my gloves and wipe them down good if I touch things like my phone. If it wasn’t for how dry iso makes my skin I’d probably not even wear gloves. I also typically do my agar work first thing after I’ve showered making sure to not pet the dogs or go outside beforehand.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: fahtster]
    #26629874 - 04/26/20 01:16 PM (3 years, 8 months ago)

There's something similar that's a biopsy tool but they are disposable and probably be hard to sterilize so would get expensive quick.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: Sockadin]
    #26629878 - 04/26/20 01:18 PM (3 years, 8 months ago)

and I prefer the flip which would be impossible with a dip n drop.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: PMBastian]
    #26630136 - 04/26/20 03:22 PM (3 years, 8 months ago)

I started with it in a SAB never had issues, just have to be mindful of hand placement and dish /lid angles.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: Hobbit GDF]
    #26630259 - 04/26/20 04:23 PM (3 years, 8 months ago)

Yea, that's why I use the cooling plate. Takes a little longer to cool down than a scalpel. Also be careful no to overheat in the first place. I flame into the side hole and out the tip instead of into front of tip for the majority that way heat not blowing toward your hand.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: redhandmat]
    #26633136 - 04/27/20 09:52 PM (3 years, 8 months ago)

Was just sent a picture of my new agar punch. Really excited, should be here in the next week or so. Can't wait to give it a try. Hopefully it works as well or better than my leather punch.



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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: Roundabout]
    #26635756 - 04/29/20 01:03 AM (3 years, 8 months ago)

Quote:

Roundabout said:
Beautiful tool and a great TEK.

My recent batch of plates had some bubbles. I had set sous vide to 120 and everything poured fine, but I wonder if it was too close to gelling temp or perhaps I swirled too much before the pour or something. Have you ever had trouble with bubbles?

In any case pulled those supplies into my cart, thanks for doing the homework on which brands make the clearest plates! Love the coloring too




Yes, I try to avoid shaking and swirling after PC as it will cause bubbles. I’d probably set the sous vide a little warmer as like you said. It would start to gel up on you toward the last few plates. I shoot for around 130f

I also avoid swirling after pc as any solids will have settled to the bottom of the media bottle and I leave the last 1/8” or so in the bottle to prevent that sediment if any from pouring into my plates.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: D3_Myc]
    #26646875 - 05/03/20 07:49 PM (3 years, 8 months ago)

Added a few more pics to TEK


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: anthocyanins] * 1
    #26649167 - 05/04/20 08:58 PM (3 years, 8 months ago)

Quote:

anthocyanins said:
Dude, v nice. So impressed.

Couple questions:

I have a very basic SAB - clear tote with two arm holes. Are you saying to leave the dishes OPEN to the air in the SAB for several days??

I guess this is more for xfers. But what about starting from scratch, just a syringe.

Any tek on incubating the dishes for pan cyan?

Thx.




Leave the lids on the dishes but don’t wrap them yet. You can cover the arm holes with press n seal or something though.

For inoculation you can put a drop to an inoculation loop. There’s great TEKs on the subject.

I haven’t grown pans yet sorry


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: anthocyanins]
    #26649180 - 05/04/20 09:06 PM (3 years, 8 months ago)

Sorry on my phone, hard to search and post teks or I’d link it for you. Start here https://www.shroomery.org/forums/showflat.php/Number/24144021


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: Grumio]
    #26656180 - 05/07/20 10:07 PM (3 years, 8 months ago)

Got my new agar punch in yesterday and gave it its first test run today. Worked really good.



Did some transfers on these plates before they went to grain. Transfers off to cold storage









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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: Crackatoa]
    #26656290 - 05/07/20 11:00 PM (3 years, 8 months ago)

Don’t want to share his personal info without his permission but yellowbirdbanjos@gmail.com is his email that he said I could share. Tell him kinoko sent ya.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: tomcards]
    #26664497 - 05/11/20 01:07 PM (3 years, 8 months ago)

Thanks


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: Sockadin]
    #26664503 - 05/11/20 01:11 PM (3 years, 8 months ago)

Quote:

Sockadin said:
Just got my punches yesterday gonna give em a test run in a bit.

Has anyone used agar punches to variety cross yet?




Was it you that brought up maybe trying to sandwich two wedges together and seeing if it would cross? Try it out! Be a fun experiment.

If anyone else is interested in the agar punches his email is yellowbirdbanjos@gmail.com  tell em kinoko sent ya.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. [Re: tomcards]
    #26664864 - 05/11/20 04:18 PM (3 years, 8 months ago)

no problem, hope it works out good for you.


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InvisibleD3_Myc
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Re: D3's Perfect transfers and agar TEK. *DELETED* [Re: D3_Myc]
    #26664953 - 05/11/20 04:47 PM (3 years, 8 months ago)

Post deleted by D3monic

Reason for deletion: oof


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