Home | Community | Message Board
You are not signed in.
Sign In New Account
Forum Index Search Posts Trusted Vendors Highlights Galleries FAQ User List Chat Store Random Growery

PlanetSpores.ca
This site includes paid links. Please support our sponsors.

Mushrooms, Mycology and Psychedelics > Mushroom Cultivation
Threaded Mode Threaded  Previous topic Previous  View all topics Index  Next topic Next 

Welcome to the Shroomery Message Board! You are experiencing a small sample of what the site has to offer. Please login or register to post messages and view our exclusive members-only content. You'll gain access to additional forums, file attachments, board customizations, encrypted private messages, and much more!

Shop: Original Sensible Seeds Bulk Cannabis Seeds   MagicBag.co All-In-One Bags That Don't Suck   Myyco.com Isolated Cubensis Liquid Culture For Sale   Sporeworks.EU Spores for European Microscopy   North Spore Cultivation Supplies   Mushroom-Hut Substrate Bags

Jump to first unread post Pages: 1 | 2 |  [ show all ]
Some of these posts are very old and might contain outdated information. You may wish to search for newer posts instead.
Tea agar myc cleaning attempt * 1
    #24562663 -

Hi guys,

Here are some pics of the petri dishes I am using for cleaning bacterial mycelium I have... Using the sandwich method

TEAA (Tea Agar) recipe used:
- 500 ml tea extract (made from 12.5 grams of green tea and 500 ml water)
- 7.5 grams of malt extract (= 50% of a regular MEA recipe)
- 9 grams of agar

Some info for the uninitiated; when you use half the nutrients (malt extract in this case) in your agar recipe the mycelium will have to search for food which makes the mycelium spread faster. This is what you want when cleaning a culture. You want the mycelium to outrun the bacteria your culture is infected with... Since tea acts as a bacterial inhibitor/anti bacterial agent I am hoping the bacteria stay in the middle of the petri dishes and the mycelium will eventually grow towards the outsides of the dishes

I will post updates/pics if things work out

Edited by Shroomway (08/18/17 12:15 PM)

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway] * 1
    #24562725 -

Awesome!  Can't wait to see your results.  I am also in the process of cleaning a a culture infected with bacteria via agar sandwich.  Just using regular agar atm, no antiboitics.  I might try your way if I don't see any improvement.

Here is my first round.  As you can see, I've got my work cut out for me.  This culture is dirty af.

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24564312 -

I hope it works out for you... The growth looks pretty nice to me... Maybe you could transfer to a tea agar plate..

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24567465 -

Update after two days of incubating at about 71F/21C. Bacteria are already visible. I am not aware of any spreading of bacteria though... No signs of mycelium growing through the upper part of the sandwich yet.. I think it is going to take a long time since the thinly poured plate for creating the upper sandwich-squares were not thin enough for my likings. I hope the myc will not suffocate... Can't wait to see if the tea really inhibits the bacteria growth. Cause the transfers are REALLY bacterial

You can clearly see some little blots of bacteria. It should not be there
since I am positive I did not touch any bacteria before I put the square
piece of tea agar onto the transferred piece of mycelium. I don't know how
bacteria ended up outside of the sandwich


Even this petri has bacteria visible along 3 out of four edges of the sandwich


This is a transfer that went wrong. The sandwich slid so you can see the imprint
of it's original position made visible through bacteria and texture change.

Edited by Shroomway (08/20/17 02:17 PM)

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24567856 -

Are the surface of your plates kinda watery?  Might be why you have bacteria growing outside of the sandwich.  Water seeped in and carried the bacteria along with it.  Just a theory.  My plates are a little watery.  Hopefully the piece on top sealed well enough with the bottom that this isn't an issue.  I made a point to keep the petris as level as possible.  We'll see.

What is you technique for extracting the mycelium from the agar?  I've been using an inoculation loop, but it's a pain in the ass.  Was wondering if there is a better way.

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24569167 -

I like using a scalpel. That's how I make my transfers. The blades I like to use have a sharp point so I can also use it to scrape very small fragments with it. For this sandwich I used tweezers to drop the square on top of the transfers. Also I find it very easy to use the point of the scalpel to pick up pieces that I cut and to drop the pieces by swiping it across the new plate.

My plates were actually rather dry. Not even condensation. But I think you are right that it might be water that carried the bacteria outside of the square. It probably happened when I pressed the square down to make a good seal. There is somehow probably enough water/moist present in the agar that squeezes out I guess... Very curious to see if the tea is going to slow the bacteria down!

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24569219 -

Cool, I'll give my scalpel a try for extraction next time.  I am also curious about this tea business.  If you don't mind, I would like to post pics of my second round of regular sandwiches on this thread for comparison against your tea sandwiches.  COuld turn into a fun, informative report for the community.

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24569259 -

WeavieWonder said:
Cool, I'll give my scalpel a try for extraction next time.  I am also curious about this tea business.  If you don't mind, I would like to post pics of my second round of regular sandwiches on this thread for comparison against your tea sandwiches.  COuld turn into a fun, informative report for the community.


Yeah, nice, please post. I like to make it as informative and think along like as possible! Thanks :smile:

I might add (not unimportant) that I was inspired by Ferather who is experimenting with T-gel

Edited by Shroomway (08/21/17 12:10 PM)

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24569284 -

Awesome!  The myc is just starting to poke through the sandwiches, might be a week or so before I have anything to report.

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24583184 -

Nine day's since the start of the transfers to TEAA (tea agar). The bacteria definitely are inhibited greatly. The plate I slipped up on (because the transfer slid) has the myc showing in two places. The mycelium is just starting to get ahead of the bacteria. I think it will be fast now because it's Pan Cyan myc. The TEAA is doing a good job!

The other transfers might be suffocating or will just take a long time to grow through the sandwich...

transfer 1). Same lighting as before for comparison with the earlyer shots


Same picture as directly above. Different light to get a look and feel for what's really going on
[/url]

transfer 2). Not poking through as earlier mentioned


transfer 3). Bacteria neatly in the same place. Almost no bacterial growth/spread

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway] * 1
    #24583945 -

What's up Shroomway, here are my results.  This round of sandwiches did not impress.  I feel my technique was improved from the first round.  There was a marginal improvement overall, but nothing to write home about.



^^This petri was the best of the bunch.  There was one edge that looked pretty good, so I took regular transfers.  The nasty shit was a good 1/2" from the edge in that one spot.  The bacteria is very persistent.  You can't see it all that well from the pics, but that shit is growing almost right up to the edge of all the cultures.  Gonna grow out the transfers and see what I end up with.  In the meantime, going to make a small batch of tea petris as a backup plan.  Anything to slow this shit down a bit.

The others:




^^Would myc that crawled up on the side of this petri be a good sample for a sandwich?


Edited by WeavieWonder (08/27/17 08:02 PM)

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24584759 -

Visually those plates are pleasing... but looks can be deceiving... I was thinking: "could it be helpful to transfer to the very edge of a plate so there is more distance for the myc to get ahead of the bacteria; making it into some kind of rat race.."?

Are you using pre made agar or half the nutrients of standard agar?

You are using fairly thick squares for the top half of the sandwich. I poured a thin plate (intended to, but still ended up too thick) to cut my squares. The thinner they are the better they "fall" around the transfer and make a seal. RR was talking about pouring it thinly so I followed his advice. Maybe someone else can comment on that. Cause thicker certainly also means a bigger barrier.. I would certainly like an expert opinion on that.

I wonder if it would be an improvement if one would make a very small cutout in the middle of the tea petri and put the transfer in it. One could put the top half of the sandwhich to cover the hole up and have less chance of contaminants squeezed out as you are trying to make a seal; you are getting an instant seal by just putting it there basically. Will be trying that for sure. One could also pour the tea agar to fill op the hole with the transfer instead of placing a square. I like this experimenting..

Maybe it IS a good idea to make a transfer from the myc that crawled up on the side of the petri. A gamble worth while maybe? There are less nutrients there for the bacteria. Maybe you get lucky and not catch a different contam..

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24585165 -

Have you taken transfers from any of your tea plates yet to grow out on regular petris?

   

Shroomway said:
Visually those plates are pleasing... but looks can be deceiving... I was thinking: "could it be helpful to transfer to the very edge of a plate so there is more distance for the myc to get ahead of the bacteria; making it into some kind of rat race.."?


They will make for very nice inoculant once I get it clean.  You can see bacteria ridden myc (grey) is riding on top of the healthy looking myc (pure white).  You can also see in places the white myc looks puffed out(stressed).  I haven't considered placing the transfer more at the edge.  I've seen pics of people doing it that way.  Maybe there's something to it.     

Quote:
Are you using pre made agar or half the nutrients of standard agar?


I was using an premix (MEA, agar agar, yeast).  30g/L (down from 50g/L) supplemented with additional agar agar.  This provided me with aggressive growth.  Couldn't quite get the agar agar dialed in.  Plates have been just a little watery despite little to no condensation.  Seems to be an issue, especially for sandwiches.

Used up the last of those, and made a new batch of petris.  This time I did the mixing.  Used 15g/L dried malt extract (1.5%), and 20g/L agar agar (2%).  The plates turned out great.  No excess water and the agar is the prefect firmness.       

Quote:
You are using fairly thick squares for the top half of the sandwich. I poured a thin plate (intended to, but still ended up too thick) to cut my squares. The thinner they are the better they "fall" around the transfer and make a seal. RR was talking about pouring it thinly so I followed his advice. Maybe someone else can comment on that. Cause thicker certainly also means a bigger barrier.. I would certainly like an expert opinion on that.


Poured the new plates a little thinner this time.  The thickness of my squares did not seem to affect penetration.  I saw myc poking through after a few days.  I can see how a thinner square would preferable.  The square would be more malleable, which would drape over the surface better, right?  My seals seemed good at glance, but evidently not.  Bacteria city.

Quote:
I wonder if it would be an improvement if one would make a very small cutout in the middle of the tea petri and put the transfer in it. One could put the top half of the sandwhich to cover the hole up and have less chance of contaminants squeezed out as you are trying to make a seal; you are getting an instant seal by just putting it there basically. Will be trying that for sure. One could also pour the tea agar to fill op the hole with the transfer instead of placing a square. I like this experimenting..


Experimenting is the best part:thumbup:.  I've tried what you describe a couple times, but didn't have any success with it.  Kinda want to try it one more time with the new plates though.  It requires an extra step, but transfering a small wedge vs a scraping myc off the agar surface is a lot easier for me.  If done correctly, I think it could work.  Gonna throw that one in the mix for my next round of sandwiches. 

Quote:
Maybe it IS a good idea to make a transfer from the myc that crawled up on the side of the petri. A gamble worth while maybe? There are less nutrients there for the bacteria. Maybe you get lucky and not catch a different contam..


It's more risky, but don't have anything to lose really.  Gonna try this next.

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24585433 -

No,..I am still waiting to make a first transfer to a regular petri or tea petri the going is very slow.. Maybe in 5 day's or so... I hope :wink:

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24585483 -

I'll Be TRYING THIS HAve A Print From OUTSIDE THAT Is Giving Me Hell Why THE Hell Is MY Phone Auto CORREVTING LIKE THIS

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24585511 -

Right on, fingers crossed.  Hope to be ready for the third round of sandwiches in a week or so.  The transfers I did SHOULD look better, but not going to hold my breath.  Going to try your tea agar recipe out this week.  Can't wait to try it!  Would you provide me with a link to the post/source you acquired the recipe from?

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24585542 -

Yup, here it is
It's not "my" recipe btw. Like you said, it has a source. It's fair to mention that it's Ferather who read about it on the internet and was the first here to experiment with this great tea ingredient. I just made it a little simpeler, left out the gelatin..

Edited by Shroomway (08/28/17 02:28 PM)

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24603609 -

Attempted to make tea agar today.  Tested the PH after making the extract and it was on the acidic side (around 6).  Not ideal.  I was instructed to shoot for a PH of 7-8.  Found some powdered calcium carbonate at the pet store to balance the PH.  The attempt was not successful.  Apparently calcium carbonate is not water soluble.  With a lot of unknowns and my limited scientific background, I decided to mothball the tea agar project for time being.  I have a lot of other projects in the works atm, and my regular life is busy as well. 

I need a sure thing so the ball can continue rolling forward.  Just got some gentamicin sulfate in the mail and whipped up a sleeve of petris.  Gonna try a new batch of sandwiches this week.  My last transfers I did were still dirty af.  I need a win here.

How's things going with you, Shroomway?

Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: WeavieWonder]
    #24605012 -

Hi there! What a pity you didn't continue with the natural experiment... Although I understand you don't have the time to experiment now. Sometimes you just need quick results and gentamicin is going to provide you with that I reckon!

I just made transfers from the plate that showed mycelium growth. By the looks of it the bacteria stayed in place. The transfers are to regular PDA and oats agar plates (no tea). Out of 4 plates I can show two pictures. The other two dishes have condensation (the oats agar dishes). Those dishes, as far as I can see, look a bit weird; wet around the transfer. Very hard to tell, it might not be bacteria but just water... Fingers crossed...

I thought the growth would accelerate once the mycelium broke free from the contams, but that didn't happen. I guess I used too much green tea because green tea is probably stronger then black tea. Next time I am likely to use maybe 8 grams of tea in stead of 12.5 grams.

The PDA dishes show no bacteria yet for sure. Good hopes for those two!. The oats agar I just can't tell... Here are some pics:


Extras: Filter Print Post Top
Re: Tea agar myc cleaning attempt [Re: Shroomway]
    #24605899 -

12.5g green tea and 7.5g DME might be a little too nutritious, which could account for your slow growth.  Perhaps cutting down a little on both next batch would provide better results.  Something like 10g green tea and 6.5g DME?  I think adjusting the PH would prove beneficial too.  In theory, a higher PH would slow down the growth rate of contams, giving the mycelium a chance to jump ahead.

I would like to pick the tea project back up this winter.  It would be nice to have an alternative to antibiotics.  All anybody needs is inhibited contam growth, not total annihilation.  Total annihilation does feel satisfying ATM though, as I've been kicking this bacteria around for months, and can't wait to be rid of it.  I might burn the spore print from which these cultures are derived.  Pretty sure someone wiped their gooch with it before sealing it.

Anyways, That's awesome man.  I'm eager to see your results.  Did my sandwiches last night so I should have some show and tell in about a week.  Good luck!

Edited by WeavieWonder (09/05/17 01:54 PM)

Extras: Filter Print Post Top
Jump to top Pages: 1 | 2 |  [ show all ]

Shop: Original Sensible Seeds Bulk Cannabis Seeds   MagicBag.co All-In-One Bags That Don't Suck   Myyco.com Isolated Cubensis Liquid Culture For Sale   Sporeworks.EU Spores for European Microscopy   North Spore Cultivation Supplies   Mushroom-Hut Substrate Bags

Mushrooms, Mycology and Psychedelics > Mushroom Cultivation
Threaded Mode Threaded  Previous topic Previous  View all topics Index  Next topic Next 

Similar ThreadsPosterViewsRepliesLast post
* G2G transfers...an endless cycle? ADoobie 7,810 18 11/12/20 07:10 PM
by FredsGotSlacks
* CLEAN Spawn transfer help! Splat 1,192 2 02/26/02 03:23 AM
by Anno
* myc transfer to grain question ksdoob 805 3 09/25/04 11:23 PM
by ksdoob
* Agar Questions Rohypnol 1,404 5 10/28/03 03:37 PM
by Mycomancer
* H2O2 use with spore syringes instead of agar? Stix Russell 1,165 3 02/20/04 07:48 PM
by Stix Russell
* grain transfer question??? beejay 1,646 12 01/10/05 04:11 AM
by MushroomFriend
* what is myc. piss? blissedout 5,973 7 12/11/04 11:50 AM
by hyphae
* clean suits and hair nets kell 1,625 15 12/16/04 07:54 AM
by kell

Extra information
You cannot start new topics / You cannot reply to topics
HTML is disabled / BBCode is enabled
Moderator: Shroomism, george castanza, RogerRabbit, veggie, mushboy, fahtster, the_chosen_one, LogicaL Chaos, 13shrooms, hamloaf, cronicr, Stipe-n Cap, Pastywhyte, bodhisatta, Tormato, Land Trout, A.k.a
1,620 topic views. 15 members, 204 guests and 112 web crawlers are browsing this forum.
[ Show Images Only | Sort by Score | Print Topic ]
Search this thread:

Copyright 1997-2026 Mind Media. Some rights reserved.

Generated in 0.043 seconds spending 0.01 seconds on 18 queries.