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Crossing strain varieties via swab to spore solution


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#1 fahtster

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Posted 10 November 2020 - 12:36 AM

So this is still a work in progress but I swabbed AA+ and PE to the same swab (had both varieties sporelating at the same time and swabbed them to the same swab—it was actually 2 swabs because my presterilized swabs come in 2 packs)

Then I made spore solution out of the swabs by rubbing the two swabs together (where there were spores) in sterile water... you would use a SAB or flowhood but I used an oven bag (That’s a whole ‘nother thread lol) and inoculated 4 pf cakes.. 3 of the cakes I birthed to a FC (the 4th I’m keeping invitro to see what it does)

F6B1EA95-813D-4A42-BCDA-E61F57EAC522.jpeg

I think my method worked! They all have bi-color and bell shaped caps

3023A4CE-30A9-4832-9651-4438CDA901F6.jpeg . 98194CFE-0512-4ABC-B918-BE0DB203E0FB.jpeg . 34B9C125-C3A9-4B1B-8903-9198344776B8.jpeg

They’re solid through and through

CCEB44D2-9597-427C-BB6F-65E9578D6D45.jpeg

They have the PE stem bumps

4FB873B5-4B09-4077-9E91-BDE5786E80FE.jpeg

Here’s the two tubs the caps for the swabs came from

AA+. D872636B-37EF-4DC4-90E4-9E1083FA0C3E.jpeg . PE... 658F7F23-ACA9-4190-A0FA-9FA5096E0802.jpeg

Comparison pics
AA+_________ PEA+
0F31E4C2-0205-40A5-B816-E552389485C5.jpeg . EB35821C-7C13-42C2-8862-A66B82BB0ED4.jpeg

They’re definitely not straight AA+ and definitely not straight PE

I think my method of getting them to cross is just really effective

1. Double swab two varieties on top of each other. The presterilized swabs I have come 2 in each pouch so I just use both at the same time and return them to the same pouch

2. Make a spore solution by rubbing the two swab tips together in sterile water.

I think the big problem with getting crosses is that spores clump and it’s hard to get the two varieties’ spores to germinate with each other but by swabbing on top of each and then rubbing the tips together, it intertwines and meshes the two varieties into the same clumps and they easily mate with each other and that’s why I think I got crosses at every inoc point.

Faht

Edited by fahtster, 10 November 2020 - 12:40 AM.

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#2 DickMoby

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Posted 10 November 2020 - 02:34 AM

Nice !

 

It definately worked.

Currently reading into cross breeding and thought about doing diluted spores which would take alot of plates and time to do.

 

This looks promising and does not collect as much spores as a double print.

 

Nice ones!

 

I'll put myself way back in the line of people for a print for when/if you feel like sharing prints


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#3 Moonless

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Posted 10 November 2020 - 03:32 AM

I like this very much! Whats the next step, are these stable as a strain and you could reproduce from spore print or dose more work need to be done?


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#4 fahtster

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Posted 10 November 2020 - 03:53 AM

Lots more work.. gotta grow out the spores and then look for variations of the two varieties that were crossed to make sure the cross happened as suspected.. then pick a fruit that has both traits like these and grow those out... do that Over and over again until you keep getting only fruits with both traits.. this would be F1.. the next grow out would be F2 after that F3 and so on.. I’d probably go to F7 before I’d even think about it being stable.. at least as far as I understand it.

This is the first time I’ve done this but it was incredibly easy and there were too many crosses in those 3 cakes to think it was a fluke. I’m putting the method out there so that everyone can try it.. hoping to make getting crosses not so difficult (you NEED agar and have to grow multiple subs in hopes of getting one cross etc)

I think part of the reason it hasn’t been done like this before is that (at least from what I’ve heard/seen) we’re always Told that you shouldn’t make spore solution from swabs because they’ll be dirty and I just don’t think that’s true.. at least not any more dirty than making spore solution from a traditional print (cap sitting on foil)

If you think about it, swabs are actually less risky because they’re sterilized and exposed to air for like 30 seconds tops (if you’re doing your swabbing correctly) and traditional prints are exposed to air for hours. One thing that I do that I haven’t seen from anyone else is that I do a prewash on my swabs before I put them in the water that I pull the SS (spore solution) from.. so I have two jelly jars of sterilized water.. the first jar is the prewash where I just dip the swab tips in them a couple times to get off any possible surface Contaminates and then move the swabs over to the second jar and dip the swabs into the water and rub the two swabs together vigorously.. that’s the jar I pull SS from.

Faht

Edited by fahtster, 10 November 2020 - 03:55 AM.

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#5 Moonless

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Posted 10 November 2020 - 04:05 AM

Nice. Is this your first rodeo making a strain? I really like the appearance of this mushroom. Its extremely distant and has distinct parents too.


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#6 fahtster

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Posted 10 November 2020 - 04:12 AM

Yeah.. first time.. thought about it for a long time (best way to go about it etc) and made the double swab last April.. just started cult again... pretty much just to play around with crossing.. next project is a red spore PE. Got that in the works too.

Faht
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#7 DickMoby

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Posted 10 November 2020 - 04:23 AM

I think especially early versions are interesting because of the different present phenos, at that stage you choose what to stabilize and maybe find something even more different during that time.

 

The PE crosses also tend to throw mutants in the unstable phase after crossing which is another interesting part imo.

 

Have you thought about a name yet ?

White Bells could be a good one 


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#8 Freaky

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Posted 10 November 2020 - 08:39 AM

Faht! This is super cool and something definitely worth trying since it's so easy. 

 

Are your cakes sitting on hockey pucks?  :thumbs_up:  I dig it!

 

I'm just learning about the new world of swabs and very into it.  I've got some APE swabs gifted to me I may try this with.


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#9 fahtster

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Posted 10 November 2020 - 11:28 AM

Haha.. they’re just those gray leak proof lids upside down and filled with wet verm.

Thanks all.

Faht

#10 Freaky

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Posted 10 November 2020 - 01:41 PM

Haha.. they’re just those gray leak proof lids upside down and filled with wet verm.

Thanks all.

Faht

 Lol - looked like right goon style pucks in there 

 

Oh hey, I got to thinking - have you considered or tried crossing the swabs on agar to mix? You explained you do a 'prewash' of sorts then rub to mix in the second sterile ss water jar.  So I'm thinking, would it be possible if you did the prewash then rubbed on top of agar? Or is it that the water suspends the spores to allow them to connect from each swab strain and agar wouldn't allow suspension or as easy of a way for them to mate between them?

 

Just thinking out loud really - but if this could be done on agar successfully and viewed under a scope it could also allow for more detailed transfers of the mating spores.



#11 fahtster

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Posted 10 November 2020 - 02:04 PM

I’m not totally sure.. I don’t use agar but I don’t see why it wouldn’t work.. only one way to find out ;). Please reply here if/when you do it. Excited to see what other ppl get with it

Faht
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#12 Freaky

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Posted 10 November 2020 - 02:31 PM

I’m not totally sure.. I don’t use agar but I don’t see why it wouldn’t work.. only one way to find out ;). Please reply here if/when you do it. Excited to see what other ppl get with it

Faht

 

I'll definitely give it a try on agar.....patiently waiting for my TC's and GT's to do something exciting lol



#13 DickMoby

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Posted 10 November 2020 - 02:55 PM

I'm also thinking about agar but not sure which varieties to choose to actually be able to recognize a real cross.

 

Those AA+ PE you can clearly see the mixture, definately gotta choose two varieties with clearly recognizable different characteristics.

 

Gave my AA+ print to a mate and don't have PE available here.

I'm currently saving a bunch of cultures that were frozen a couple times, some look pretty good, just put them all to grain yesterday hoping they produce fruits that mature, the frozen TAT culture aborted at about 2cm height, went back to an older plate that was also frozen and cloned a pin from it which gave me some good rhyzo growth, maybe that fruits better.

 

If one of them produces spores: TAT, Albino Cambo, Albino Burma, Kape and Rusty Whyte then I could try to cross something with an albino, maybe Melmac x A Cambo, Melmac x TAT or Melmac x Rusty Whyte, that could be sexy.

 

Faht this post has me a little exited , I can't afford a microscope right now but would want to aswell give that a try very soon.


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#14 fahtster

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Posted 11 November 2020 - 03:07 AM

Just now I came up with a way to very quickly get layer upon layer of 2 different varieties onto one swab (I use two because I like to rub them together when making spore solution—which further clumps them together)

made a diagram :)

592EB633-3AB1-4CB5-807C-EBEDCF505613.jpeg

Gonna test it out tomorrow with two caps from a mini mono that’ll be ready.. just to see if it’s a pain in the ass to do.. probably easiest if they’re the same sized caps.

Thinking that it’d be even cleaner if at first I take one pair of swabs and run them a couple times over both gills (where they meet) and then set those aside.. grab a new pair of swabs and do the back and forth (like 10 times) over the same gills.. the first set of swabs will act as a sort of cleaner and get any contaminates that are sitting on the surface.. I’m sure they’ll still be plenty of spores for the 2nd (the ones that count) set of spores to grab

When I made the PEA+ pf cakes, I also inoculated a jelly jar of rye and once that was colonized, I made GLC and inoculated 2 qts for a mini mono. Here’s what the jelly jars look like

5B6612D5-0257-4761-B785-2C5BB6B881A7.jpeg

After I used the GLC on the qts, I actually took the jelly jar of grains and put it to CV in a little glad container and ziplock (needle holes poked in it every 2-3”) with a little wet perlite and it grew out 2.5 dry grams.. nice tester dose



6B5DE75E-C663-4772-89AA-5AE3537A4CBC.jpeg . 765F9F17-65B6-4C34-B39C-A6B51578368E.jpeg . CB1E343F-BD6A-4F0B-8ED0-CC950B1F8D3B.jpeg

Here’s the PEA+ mini mono today (I cased half of it just in case if needed it because if the PE part of PEA+)

93B7FA18-1283-4A0F-A5DC-CADA8D38FECB.jpeg

I pulled the big guy in the middle and it’s solid like a rock

Faht
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#15 DickMoby

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Posted 11 November 2020 - 08:09 AM

I like boobs  :ph34r:

 

The pre wipe sounds like a good idea, for the cut I'd think it could maybe introduce some dirt from outside of the cap into the gills.

 

Did it throw some more of the ones from your very first cake ?

Those white caps with big brown nipples were great.

 

Something about the PEs.

Do only ~5% drop spores or do only those 5 produce spores at all ?


Edited by DickMoby, 11 November 2020 - 12:19 PM.

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#16 fahtster

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Posted 11 November 2020 - 11:46 AM

The cut is straight down (no sawing action) once with gills facing up so the blade only touches the gills. I tried it this morning.. worked great but I had to cut a flat piece on the top of the caps first

730290B6-69C9-4614-ACED-DE8A8FAD613B.jpeg

Then they sat flat next to each other.. definitely gonna work better with bigger caps

6BC33DE4-C411-4A0D-9050-496E2322FB57.jpeg

Here’s that tub today

6658D521-E212-47CE-B395-F6EB4EC2D2C2.jpeg

All PE make spores.

The tubs phenos aren’t as pronounced as the cakes but they’re definitely PEA+.. I expected that the cakes would have more since I shake the GLC grains for the tub.. more chance of 1 or 2 sub strains to overpower all the rest unlike cakes that don’t get shook

Faht
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#17 DickMoby

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Posted 11 November 2020 - 12:52 PM

Get it.

 

Thought about cutting the cap gills down which would bring some nasties into the gills but sure gills up it wouldn't be an issue.

 

Beginning of this year I put an rtv port into one of these micro boxes

 

PSX_20201111_183054.jpg

 

Maybe for the pheno hunt somthing like that could be a good thing, the box was just inoculated and then cased, thats where the dirt water comes from.

 

No mixing after the inoc and a flat surface for mushs to grow on.

 

Looking forward to see where the PEA+ journey goes


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#18 fahtster

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Posted 11 November 2020 - 03:00 PM

Yeah for sure.. another OMC member called Cronicr uses the rectangle glad containers with inoc ports arranged all over the edge of it, makes large pf type cakes to do crossing and pheno hunting and it works well.

Faht

#19 fahtster

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Posted 11 November 2020 - 09:42 PM

Last money shots of the PEA+ mini mono (I’ll post dry weight tomorrow.. it’s in the dehydrator)

3F67936D-ADA0-4969-9A19-766B89153E3B.jpeg . AC8847AC-FEDA-4F73-855A-5B4D704B924F.jpeg
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Took a bunch of swabs (still have the culture too)

BE519A09-1859-47A6-B839-8552EBC5191F.jpeg

Those 3 pf cakes’ fruits dried down to 16.1gms (5.35gms per cake—not too bad for MS)

0F815605-D973-48EB-87FC-610670D32DCF.jpeg

Faht
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#20 DickMoby

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Posted 12 November 2020 - 07:42 AM

The one in the dehydrator gives me strong Melmac vibes, nice!

 

Do you sell swabs ?

Could Paypal a small donation  






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