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getting an isolate from a strain
    #6550353 -

Hi, i have been looking for a good tek on how to get an isolate of a strain on agar plates. I havent had any luck finding a tek, so i turn to the infinatly knowledgable community.

any info is very much appreciated. if it makes a difference, i'm planning on using Potato dextrose agar.

thanx,
synergy


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Re: getting an isolate from a strain [Re: shroomsynergy]
    #6551349 -

Hey, I'm interested in this as well.

This link was posted on another thread about isolation: http://forums.mycotopia.net/showthread.php?t=8303


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5 shroom me!

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Re: getting an isolate from a strain [Re: shroomsynergy]
    #6553624 -

http://www.shroomery.org/8530/Spawn-Spawning-and-Spawn-Growth



From the above link.


Quote:
Spore Cultures
Mushroom spores are found on basidia lining the gills of the cap. To collect the spore, the cap is hung over sterile filter paper in a spore collecting chamber. Spore cultures differ genetically from tissue cultures. Cultures originating from spore germination are unpredictable. Spores germinate creating many different genetic combinations that are desirable when attempting to develop new strains, but undesirable when trying to maintain a specific strain. There is no way to predict if a culture's fruiting potential is similar to its parent other that to test many different lines obtained from spore germinations. Hundreds many be tested before obtaining a culture with good production potential.

Culture Validation Cropping Trials
There is no in-vitro test to determine a stock culture's validity. A series of cropping trials must be conducted on the mycelial stock culture to determine a culture line's value. Mushroom yield, size, color, cap shape and any other desired quality or growth factors are selected and then compared for each culture line.



Put some spores on an agar when they grow cut a piece of agar from your plate and put it on another agar plate. That is your isolate. Until you get it to fruit though you can't tell what you are working with.

cropping trials must be conducted on the mycelial stock culture to determine a culture line's value

Once you have some isolates, they need to be tested for their ability to fruit.

Edited by falcon (02/11/07 12:11 PM)

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Re: getting an isolate from a strain [Re: falcon]
    #6554041 -

So, it would be better to get a live culture from fruit, that way you have an idea of what to expect?

Makes perfect sense.


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5 shroom me!

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Re: getting an isolate from a strain [Re: Jorsher]
    #6554234 -

So, it would be better to get a live culture from fruit, that way you have an idea of what to expect?

Ummm, that is one way, you could clone one of the fruits from an isolate that you have grown. Keeping the culture you start with on agar is the better approach.

To test an isolate transfer some of the isolate to grain or whatever to see how it will fruit, but also keep the culture going on agar. Since whatever fruits will have the same genetics as whatever is kept on agar it will be the same isolate. This is how it is done ideally.

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Re: getting an isolate from a strain [Re: falcon] * 1
    #6554819 -

The point to remember is a single swipe of spores in agar will deliver up to 100 isolates. I make the first transfers within a couple of days of visible growth on the petri dish. I'll make at least ten transfers from the perimeter, or leading edge of the mycelium. Each transfer should be no larger than a grain of rice or uncooked grain.

After a few days, each of these transfers will have grown out enough that you 'might' be able to see some sectoring. a sector is a growth that is separating itself from the other growths on the plates. They will radiate out from the center like spokes on a wheel. Transfer each of these growths to a new plate. When you have completely isolated a strain, the mycelium will homogeneous like in the 10 o'clock to 3 o'clock position in the bottom section of the picture below. The other sections have similar isolates, but all need at least one more transfer to be truly homogeneous monocultures. The petri dish below represents the fourth transfer of it's mycelium line from the original swipe of spores. Other mycelial lines were being generated at the same time.

Remember, you don't isolate ONE strain. You isolate ALL of the rhizomorphic strains from that one swipe of spores and fruit out each isolate separately to determine which has the qualities you're looking for. While you're fruiting them out, the original petri dish from each isolate gets labeled and goes into the refrigerator until you've determined which ones are keepers and which ones are trashers. The keepers are transferred to test tube slants as soon as possible, where they can be stored under refrigeration for years without substantial degradation.

I've filmed a complete strain isolation tek for my video that is coming soon, so you'll be able to see the whole process from mixing the agar to the finished isolate in finite detail, including master slant preparation and inoculation.
RR



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Download Let's Grow Mushrooms



semper in excretia sumus solim profundum variat

"I've never had a failed experiment.  I've only discovered 10,000 methods which do not work."
Thomas Edison

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Re: getting an isolate from a strain [Re: RogerRabbit]
    #6555300 -

thats alot of good advice and tips!!! thanx a bunch people!!!

You gotta let us all know when you get that video out roger!!!!! that will be badass i bet!!! are you going to sell them???


...also, were do i get dirt cheap petri dishes?? links anyone??


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Re: getting an isolate from a strain [Re: shroomsynergy]
    #6561856 -

RR, that portion of the petri dish is HOT! I have a few strains on the way thanks to spores101 and also have some potato dextrose agar and canceled my order for the glass petri dishes to get the plastic ones like you said :smile:

I'll admit I'm one of those punkass kids that downloads stuff from bittorrent, but when your dvd is released I'll be the first one to buy it, even if it appears online (which unfortunately I'm sure it will :frown:).  Any estimates on when it'll be out?

ShroomSynergy, http://www.mycofactory.com/contents/en-us/d54.html you can get 60 for $18 + shipping.


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5 shroom me!

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Re: getting an isolate from a strain [Re: Jorsher]
    #6562430 -

I won't give a date, but filming is complete and it's in post. It will be worth the wait, i promise. I doubt much of it will be online, because it's a 2 DVD set, so it would use a lot of bandwidth. There's also a lot of cool special effects that won't be visible when it's compressed for posting, so I'm not worried. Besides, I'll sue the shit out of any website or server farm that hosts it.
RR


--------------------
Download Let's Grow Mushrooms



semper in excretia sumus solim profundum variat

"I've never had a failed experiment.  I've only discovered 10,000 methods which do not work."
Thomas Edison

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Re: getting an isolate from a strain [Re: RogerRabbit]
    #6562452 -

Bittorrent is the problem though. I've downloaded 16gb HD rips of movies. People don't care about size anymore. We got a 7mbps connection and it's not even the best in our area... And it's hard to sue them since it's bittorrent, it's not hosted by any server. I'm not promoting it, just being honest. But I hope the best for you! I will definitely purchase it the day it's out, and I'm sure most of shroomery will follow. Looking forward to it!


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5 shroom me!

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Re: getting an isolate from a strain [Re: Jorsher]
    #6564226 -

hey Jorsher, what strains did they hook you up with??? did they tell you?? I'm very currious.


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Re: getting an isolate from a strain [Re: shroomsynergy]
    #6564880 -

I got to choose.  I bought orissa and cambodian the day before (i think, if not then I will soon) and I got brazilian, south american, and treasure coast from the contest.  Will have to dust off the ol' microscope!

Will probably send to my friend to do isolates of orissa just to see how big of a shroom she can grow.  The others will be for trying to get a good crop :smile:  Really excited!

Thanks again spores101!


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5 shroom me!

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Re: getting an isolate from a strain [Re: Jorsher]
    #6565297 -

those OI can get huge-gantic just by multispore inoc!!! potent too.
my fav. right now is the TC, the ones that i saw had caps kinda like mini Aminita Muscaria!!! i heard that they are kick ass for isolating, as there are alot of variations like albino, mosters, and off colors!!!

have fun checking out those under the scope!!!


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Re: getting an isolate from a strain [Re: shroomsynergy]
    #6565304 -

Yes I think I will enjoy studying them. Maybe I can share some of the spores with you in the future, or trade for others.


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5 shroom me!

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Re: getting an isolate from a strain [Re: Jorsher]
    #6566418 -

sure, let me know,  i have quite a few different prints i'd trade ya!:mushroom2:
I love my microscopy!:grin:


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