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Agar Work


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#21 CatsAndBats

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Posted 25 November 2014 - 09:59 AM


 

*Archive Material*

What is the meaning of this? Isnt everything archived? Whatever it means, i am very humble that something i contributed is going to be available for a long, long, time..... if that is what *Archive Material* represents anyway.

I think refers to the locked up stuff used as encyclopedic reference for topians. 


Edited by catattack, 25 November 2014 - 10:09 AM.

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#22 Microbe

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Posted 25 November 2014 - 12:18 PM

attachicon.gif20141125_094129.jpgattachicon.gif20141125_094152.jpg
Can take more. I use para film..
Looks like one tly shows growth and a b plus as well.
The b plus is a newer print so probly started faster due to this
When i say growth i just meen tiny whisp that u can only see at an angle so no pics

Black food coloring helps. ;)
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#23 Microbe

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Posted 25 November 2014 - 12:46 PM

My playes that were smeared are starting to show signs of life, not worthy of a pic and the EC plates have recovered and the agar plug is compleltly colonized and should start growing out across the surface. I expect it to be weak, slow, and requiring several transfers before i feed it some oats but we will see ;)

Edited by Microbe77, 25 November 2014 - 12:51 PM.

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#24 Microbe

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Posted 25 November 2014 - 12:49 PM

AGAR EXPERIMENT UPDATED.

https://mycotopia.ne...r-experimenting

#25 torn2bits

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Posted 26 November 2014 - 12:49 AM

Microbe, well written and explained well.

Agar is very intemidating for many people including myself.

I think agar to be a "test plane" if that makes sence...

1 Test new strains.
2 Testing for contamination
3 Long term Storage of culture

Stainless isolation IMHO would be best started from sterile fruit material,which would cut at minimum 50% of the process of obtaining a isolate.

Great work,it took a TON OF BEGGING to coax you to to do this thread,I the end it will be invaluable to Many.
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#26 Microbe

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Posted 26 November 2014 - 10:03 AM

Microbe, well written and explained well.

Agar is very intemidating for many people including myself.

I think agar to be a "test plane" if that makes sence...

1 Test new strains.
2 Testing for contamination
3 Long term Storage of culture

Stainless isolation IMHO would be best started from sterile fruit material,which would cut at minimum 50% of the process of obtaining a isolate.

Great work,it took a TON OF BEGGING to coax you to to do this thread,I the end it will be invaluable to Many.

I agree that isolating from a clone is where it is at for sure but after a few transfers it is close then might as well continue it out. I always make at least transfers and like to get rhizo growth before i put it in any jars. Not isolating but just selecting a good culture. Im going to isolate my culture 1-1 from my agar experiment thread as it is very close. Now this wasnt just recently started on agar. This had an oppurtunity to grow out across grain surface via 2 g2g transfers and I believe mycelium pretty much isolated itself for me, if you were to take the 3 agar plates that the myc grew out on, combine it with the 2 complety full grain jars and lay it out, thats alot of surface area. I am starting to think that my little plates are great for germinating, testing, and even increasing inoculate but for isolating i need to get some larger ones which will reduce the stress caused during the transfer and if i remember correctly every time you conduct a transfer it becomes the next generation. So for simple conversation it would be better to use 2 large plates with only 1 transfer versus 8 plates with 7 transfers. At least i think so anyway lol.....
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#27 Microbe

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Posted 27 November 2014 - 10:20 AM

AGAR EXPERIMENT FINAL UPDATE

https://mycotopia.ne...r-experimenting

#28 Microbe

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Posted 27 November 2014 - 10:33 AM

EC PLATE UPDATE

Cultures still would not getting grain fed meal from me. But i hooing that by the time it grows out and then i make my second transfer and grows out again, it will be ready.

4a4c0a1a7e024e9fe801772ce9301aa2.jpg

ca4c60080b9b53b0c315f795caec32a1.jpg

THE SMEARED PLATES

I dropped one plate and it shattered and the other started getting bacteria on it and i wasnt messing with that because i know from experience this B+ strain did not do well with bacteria contamination. But i have 2 plates that will inoculate 4 quart jars each and then g2g to expand from there if i needed. This growth should be very thick and Rhizomorphic as it has in the past and this is the true value in isolating cultures rather it be a isolate or not and using master slants.

122e35fadd0ca311c5f179e08020756e.jpg

1742dbda6d6f9d8eba147c52565f3f25.jpg

This is as far as dare to venture in this hobby today as the family functions are about to begin that is my priority today. HAPPY THANKSGIVING MYCTOPIA, enjoy this time with family and friends but more importantly be safe.
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#29 GrowerOfShrooms

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Posted 27 November 2014 - 12:02 PM

Keep up the good work bud, happy turkey day
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#30 invisibilitysyndrome

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Posted 29 November 2014 - 08:57 AM

Hey 77
so in refference to my time frame question.
ive not seen growth on the spore to agar plates. But its only been 8 days.
when do i call it bust? 20 days? I wonder what the %of bunk transfers is...i mean i did 20 so somthing gotta grow...
ive got growth on a couple that were newer spores but i cant tell if its contam or fungus

id deff say that ill make a solution first next time i do plates...this spore to plate is too long. Ive got patience and itll prob be easier to wait next time but waiting 15 days to see growth is a bummer...not to mention im handleing them everyday. Im bound to fk em up befor too long i keep touching them. I need another incubator cause my spawn jars and bags are all, in the same tote and its gotten crouded..15 lb wbs bags 20 jars and 20 plates is recipe for disaster..
anyways off topic...i can shoot pics if u think u can id

#31 Microbe

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Posted 29 November 2014 - 11:42 AM

.

Edited by Microbe77, 29 November 2014 - 12:00 PM.


#32 Microbe

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Posted 29 November 2014 - 11:50 AM

SMEARED PLATE UPDATE

This is why a created a master slant because this is the growth that was seen in many of my jars, tubs, and fruited nicely . But if you look at the 2nd pic you can tell that it is not an isolate. My point is master slants are not just for preserving monocultures. I will go ahead and isolate before putting back into a master slant.

3d0506b45cc37fd16e554b57a1911bce.jpg

c7a2d7ecacce51be2e651daf0f2dceff.jpg

EC Plate

1st transfer /second plate. These plates still are not anything i would put in my jars. Just wanted to share the current result and this is why i just dont inoculate jars with MS. Typicaly after the second transfer/3rd plate i will have a decent culture.

a5b3a08db8699fef2eae068784d294e7.jpg

My favorite thing to do right now is agar>2 transfers >grain>g2g>agar. This gives the myc 2 grain jars to grow out in to continue to develop and isolate itself. Im not saying this gets you an isolate but does a lot of the work for you.

Edited by Microbe77, 29 November 2014 - 11:58 AM.


#33 Microbe

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Posted 29 November 2014 - 12:00 PM

Hey 77
so in refference to my time frame question.
ive not seen growth on the spore to agar plates. But its only been 8 days.
when do i call it bust? 20 days? I wonder what the %of bunk transfers is...i mean i did 20 so somthing gotta grow...
ive got growth on a couple that were newer spores but i cant tell if its contam or fungus

id deff say that ill make a solution first next time i do plates...this spore to plate is too long. Ive got patience and itll prob be easier to wait next time but waiting 15 days to see growth is a bummer...not to mention im handleing them everyday. Im bound to fk em up befor too long i keep touching them. I need another incubator cause my spawn jars and bags are all, in the same tote and its gotten crouded..15 lb wbs bags 20 jars and 20 plates is recipe for disaster..
anyways off topic...i can shoot pics if u think u can id

There are so many variables that determine these time frames. How old were the spores is this unknown? I have had spores take 3-4 weeks to germ. So when to call it a bust? Thats subjective and would say more then 2 weeks tops for me. Not that means the plate is bad but the process isnt quick enough for me. Spore to agar probably gives the best chance at germinating a clean culture and that is the only reason i see doing it IMO. There is no clean print, i dont care how careful you are as the fruiting process is not sterile and making spore syringes or MS solutions you are playing Russian Roulette. Lets say you put an entire spore print into a 60 cc syringe, you dispense the solution a single drop at a time, you will dispense a contamination because i gaurantee you they are in every syringe and every solution. Its the concentration of the contamination that makes either viable or not. Im off track also and i do that alot so you and i are going to het along great!

I would give your playe at least 2 more weeks. If there is no signs of contaminations, which they are very easy to spot on agar, then i would think that the spore isnt hydrating. When inoculating a plate with dry spore then i would lower the agar ratio which helps the agar to remain a little more moist on the surface helping with hydration. If you used a premix then you.would not be able to do this without throwing off your nutrient ratios. Incubation is important for germination in my opinion. After that you could leave in room temperature.

Can you tell me about your agar? Type, ratio, and etc?

Inoculation method such as loop, scalpel, in a SAB or in front of a flow hood. And for sure get some pics as that always helps.
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#34 papa_legba

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Posted 29 November 2014 - 02:42 PM

good info in this thread. pulling up a chair :thumbs_up:



#35 Microbe

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Posted 30 November 2014 - 01:12 PM

Update on B+ smear

Looking very nice and i may have already mentioned it above this will be the one that i hopefully take to a true monoculture. So i have 2 plates of 2 diffrent strains that are ready and close to being an isolate or monoculture, at least i hope.

94257913d7ebd9362406f58b12aae6b6.jpg

How many sectors do you see?
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#36 torn2bits

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Posted 30 November 2014 - 02:38 PM

Man I man that's just beautiful
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#37 torn2bits

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Posted 30 November 2014 - 02:44 PM

The b+ on agar this is the B+ culture that you started from spore is it not.
Same .B+ you've been working on?
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#38 Microbe

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Posted 30 November 2014 - 02:54 PM

The b+ on agar this is the B+ culture that you started from spore is it not.
Same .B+ you've been working on?

Yeah. My very first ever spore syringe i purchased a year ago. I started it on agar, transferred a few times, put into grain, then i tool a sample of colonized sub and put it back on agar and transferred out and finaly put in a master slant. Im going to fruit a tub for printing only and Hopefuly i can get a few dozen prints out to myctopia. Its crazy its already been a year and im still using the same culture. Thats the beauty of master slants. A single spore print will last me a lifetime.

#39 Microbe

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Posted 30 November 2014 - 03:03 PM

I transferred to a few fresh plates an into a sugar LC, yes you read that correctly a sugar LC even after my opinion on them. Im just thinking of an addition way to store a master culture for immediate use other then in a master. I will still only use g2g or GLC to inoculate jars i attend on fruiting. I will use the lc to inoculate plates or jars i attend on using to make GLC.

I realized i could record a video and then review and screen shot to capture a image however the video stopped at some point and only captured the contaminated plate im trying to isolate from. The video may help new people to agar work as far as process not really so much about myc selection. How can i post a video that does take someone to an external link?

#40 GrowerOfShrooms

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Posted 30 November 2014 - 03:17 PM

Same way you post pictures. I use the Tapatalk app




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