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Offlineoytothevey
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Cloning, senescence, eternal master slants, and more
    #26434097 - 01/15/20 12:16 AM (5 months, 25 days ago)

So, don't hate me, but I have a lot of questions, and YouTube and going through forum posts are no longer cutting it. I'm attempting to grow morels by starting lots of grain bags and scattering them in a woods i know. So here are the questions, also I really hope this is the right place for these questions- if not delete it

If I clone a mushroom from a scraped off bit of its innards on agar, will that then (assuming no contaminants) be an isolated strain? Or will it require agar plate transfers for strain isolation?

The cloned mushroom assuming it is an isolated strain using the above scraping off a bit of its innards and wiping in a agar plate method, will it then grow mushrooms identical to the original cloned mushroom?

Could I then clone those mushrooms for a eternal supply of that specific mushroom strain?

How long can you transfer from a master slant? What do you do when it starts to dry up or run out of mycelium? From what I've read you cant keep transferring from a master slant to make more and then transferring them to make more or something bad happens with the genetics...?

What is senescence?

Thanks, hopefully this is not too much at once...


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OfflineKmacmo
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Re: Cloning, senescence, eternal master slants, and more [Re: oytothevey]
    #26434239 - 01/15/20 02:45 AM (5 months, 25 days ago)

I'm not sure if morel mycelium is much different than cubensis, but if you clone a fruit to some plates they will be pure copy's of the original sub-strain of that fruit.. The more you transfer you could loose some genetics.

Master slants can last upto couple years if you don't ever use any of it, once your running out of mycelium in the slant transfer a piece to a new master slant and just keep doing that.

Senescence is something to do with the mycelium growing and loosing strength/viability, in a master slant in the fridge the mycelium growth is basically halted meaning senescence will not be a issue.

Hopefully that helped but I'm sure someone with alot more knowledge could weigh in on this as I'm also keen to learn more :smile:


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OnlineMLPismyOPSEC
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Re: Cloning, senescence, eternal master slants, and more [Re: oytothevey]
    #26442762 - 01/20/20 09:59 AM (5 months, 19 days ago)

Quote:

oytothevey said:
If I clone a mushroom from a scraped off bit of its innards on agar, will that then (assuming no contaminants) be an isolated strain? No, it will simply be a slimmed down gene pool. Or will it require agar plate transfers for strain isolation? Correct. Many, many transfers until isolation. Talking 50+.

The cloned mushroom assuming it is an isolated strain using the above scraping off a bit of its innards and wiping in a agar plate method, will it then grow mushrooms identical to the original cloned mushroom? Yes and no. Cloning mushrooms doesn't work how cloning plants works, it's more like humans having babies. Two 6' tall, blonde hair, blue eyed parents COULD have children that are also 6', blonde hair, blue eyed, but it's not a guarantee. Percentage chance is high, but it's not absolute.

Could I then clone those mushrooms for a eternal supply of that specific mushroom strain? Yes and no. You wouldn't want to clone a clone, then clone that clone, etc ad infinitum because each successive clone slims down the gene pool even further. Some of that gene pool includes vigor/growth rate. Multiple clones of clones will eventually slow down, they will start to mutate again and not produce similar looking fruits. That's (a quick and dirty overview of) senescence. You'd want to keep the colony alive on agar and expand through spawn expansion. G2G, LC, etc.

How long can you transfer from a master slant? What do you do when it starts to dry up or run out of mycelium? From what I've read you cant keep transferring from a master slant to make more and then transferring them to make more or something bad happens with the genetics...? The mycelium won't run out, but the slant drying out is a concern. Properly made slants in culture tubes last years. An agar plate poured thick will last up to a year. Transferring A2A will effectively never reach senescence. Senescence is really only a concern when you're expanding spawn too far. If you make one quart jar of spawn, and G2G that to 12 more jars, then G2G those to 12 more each, and so on, THAT'S how you'll reach senescence. The first couple transfers will be fine, the later transfers will start to slow down and break down. I don't have a good reference on how much you can stretch it though.




Sounds like you've done a lot of research though! You're on a good track my friend :thumbup:


Edited by MLPismyOPSEC (01/20/20 10:02 AM)


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Offlineoytothevey
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26449914 - 01/24/20 02:27 PM (5 months, 15 days ago)

Wow thanks for all of the information, I have been really busy and have not had time to check- thank you!


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Offlineoytothevey
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26449923 - 01/24/20 02:31 PM (5 months, 15 days ago)

Quick follow up question,

Are you saying if I have a isolated or semi-isolated strain in a slant, if I wanted to keep that strain forever, I could just transfer some of that slant to another slant when the original begins to dry out? How do the pros keep a good strain alive?


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Offlineoytothevey
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Re: Cloning, senescence, eternal master slants, and more [Re: Kmacmo]
    #26449929 - 01/24/20 02:32 PM (5 months, 15 days ago)

Thank you for the information! Im basically trying to figure out how to isolate a good strain and that when I do, keep it alive for as long as I need it.


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OfflineDigitalRhizae
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Re: Cloning, senescence, eternal master slants, and more [Re: oytothevey]
    #26450342 - 01/24/20 06:50 PM (5 months, 15 days ago)

That's essentially what the professionals do. But for real longevity their storage conditions use liquid nitrogen to keep the cultures cold and stable. I've read that by doing it this way they can have a master slant for 10+ years before needing to make new slants. And by taking a very small portion from the master slant and expanding it onto agar, which can further be expanded on agar for a few generations and then those can be further expanded to grain, which can also be expanded several times before going onto fruiting substrate you can keep that same culture running for many years.

I remember reading that it's a 200:1 ratio before you start to run into problems. If I understand this correctly 1 liter of grain spawn, for example can be expanded to 200 liters of grain spawn.


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OfflineKmacmo
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Re: Cloning, senescence, eternal master slants, and more [Re: DigitalRhizae]
    #26451462 - 01/25/20 11:00 AM (5 months, 14 days ago)

Quote:

creamcorn said:
working from a spore print starts all over again.  two monokaryotic mycelia meet, and "mate" and you've got a fresh set of genetics.

using mycelium to inoculate more mycelium (as in an agar transfer, a gran to grain transfer, knocking up an LC from an LC, "cloning a clone") etc is the generation thing, and can be done a couple of times but when you overdo it you can run into problems.

we're not talking about generations in the traditional sense, where you think of kids and parents and grandparents and such... its more or less how many generations we're keeping the SAME set of genetics alive.





Quote:

AyePlus said:
Ideal circumstance
Master plate gets made into master slant, at the same time 2nd gen master plate is made from the master plate to be used for inoculating and the master plate and slant go into cold storage. Once the 2nd gen master is gone a new plate can be made from the og master until that one is gone, then at that point you make a new master plate from the slant and continue on like that till the slant is almost gone then you make a new slant from the last master plate.

Keep the genetics as young as possible. Cold storage slows the aging process almost to a stop but after ~ 5 years slants should be plated and grown out and re-slanted, plates redone every 4 months or so.




https://www.shroomery.org/forums/showflat.php/Number/8468463/fpart/all/vc/1
RR's notes on cultivation:
*It was ANNO that pointed out - young vigorous mycelium does better than old. However, I have gone out 5 G2G transfer generations - without ill effect. Genetics plays a part in it. In nature - mycelium is designed to grow out & fruit in one season. You can FOOL with Mother Nature - but extending it to far out - usually brings on some mutation. G2G GENERATIONS

Some good advice I got for my post that was similar to this a while ago and RR's notes are a great read if you get the time :laugh:


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InvisibleBabuFrik
Logs, wood chips, guerilla beds

Registered: 01/28/20
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26458765 - 01/29/20 06:14 PM (5 months, 10 days ago)

MLPismyOPSEC: that's pretty cool info about cloning and how that ties into selection/genetics. Do you remember where you picked all of that up from? I'm working my way through Watkinson, Boddy, & Money's "The Fungi" right now, but I'm always on the lookout for more cool material like that.


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OnlineMLPismyOPSEC
That One Ponyfucker
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Re: Cloning, senescence, eternal master slants, and more [Re: BabuFrik]
    #26460493 - 01/30/20 05:30 PM (5 months, 9 days ago)

Lots and lots of reading and inferring answers from around the Shroomery! As i was getting into agar, i saw topics of cloning and such and that piqued my curiosity even though i was a total beginner (i still consider myself pretty fresh, a year in). I wanted that full canopy like all the pros here! I'm running my first clones now, spawned them a week ago so we will see if i've achieved my first canopy.

Also Babu was great!


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InvisibleBabuFrik
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26461553 - 01/31/20 07:54 AM (5 months, 9 days ago)

Yeah same here. I've been lurking around this place for years. It's still difficult to find good literature that explain things well even after entering academia. Then again I'm also inundated with other reading so maybe I'm just not looking hard enough. If you come across something cool in your shroomery travels and think of it, drop me a line.

Babu's dialogue was hilarious right!?! Even though he looks like a wrinkly old man butt, he pretty much carried that whole movie for me. At the theater, the crowd was loudest when he popped up in the cockpit and shouted "hey heyyy!" I changed my calendar alert sound to that and it brings me tremendous joy every time I hear it.


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OfflineDr3
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26663139 - 05/10/20 09:57 PM (2 months, 1 day ago)

So there is something I've been wondering. I have a few clones running on agar plates, and I've tansferred those a couple of times using tiny wedges.

You say in your post that cloning a clone narrows the genetic pool. But if I'm transferring A2A, Im narrowing the number of strains I'm working with each time. Could this also have the same ill-effect?

Im assuming here I'm not aiming for or obtaining and isolate.

Thanks.


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OnlineMLPismyOPSEC
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Re: Cloning, senescence, eternal master slants, and more [Re: Dr3]
    #26666052 - 05/12/20 10:48 AM (1 month, 30 days ago)

What i mean by cloning a clone is now that you have your culture from your cloned fruit, if you fruit that new culture, then take another clone from that round, and so on, that you could run into issues.

Starting with parent plate A, the first plate you streaked spores on. Taking A to grain, then spawning, and grabbing a fruit to clone. Putting that clone tissue on agar makes plate B. If you take plate B all the way through fruiting, and clone that, now you have plate C. Do this enough times, and you'll run into problems.

It's unclear how many times you can do this before issues arise (and i'm confident it won't happen within the first half dozen at least), and realistically, if you keep plate B or C around as a master and always pull a wedge from there to fruit, then you won't have any problems.

Short answer; unless you're expanding grain multiple magnitudes, it's a non-issue. :thumbup:


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OfflineDr3
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26671221 - 05/14/20 08:42 PM (1 month, 28 days ago)

Thank you for your answer. I'm not talking about cloning a clone though. I am referring to the fact that whenever I make two working plates (B and C) from a master agar plate (A) , these plates will not be identical, since we are not working with an isolate.

Having taken those minuscule wedges from different areas of the plate, the strain composition of (B) will not be identical as that of plate (C). Therefore it stands to reason that they will perform slightly differently than the master plate, and also between themselves.

I understand cloning a clone serves no purpose, generally. I'm asking about the above procedure.

Thanks for your input!


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OnlineMLPismyOPSEC
That One Ponyfucker
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Re: Cloning, senescence, eternal master slants, and more [Re: Dr3] * 1
    #26714861 - 06/02/20 08:50 PM (1 month, 9 days ago)

Sorry it took so long to respond, it's been a minute since i was online! Again, short answer; don't even worry about it. Long answer; you may eventually run into senescence, but it would be many, many transfers down the road. Easily 50+ from the parent plate.


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OfflineBsdgaou
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26723211 - 06/06/20 03:23 AM (1 month, 6 days ago)

Is "senescence" thing in agar to agar transfers?


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OnlineMLPismyOPSEC
That One Ponyfucker
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Re: Cloning, senescence, eternal master slants, and more [Re: MLPismyOPSEC]
    #26726775 - 06/07/20 04:55 PM (1 month, 4 days ago)

See my last post.


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Mushrooms, Mycology and Psychedelics >> Gourmet and Medicinal Mushrooms

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