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Mushrooms, Mycology and Psychedelics > Advanced Mycology
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Mushroom genome
    #2152154 -

Just wondering if anyone has mapped the mushroom's genome sufficiently to identify the genes responsible for producing psilosin and psilocybin? We need a good winter project, and splicing these genes into an easier to grow organism sounds like a fun challenge.
Any info would be appreciated as mapping the genome ourselves could take quite a while.
thanks

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Re: Mushroom genome [Re: rwpride]
    #2152215 -

Although I really like the idea, Unless you have access to some very nice equipment, I don't think we will be messing with this for a long long time...


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Re: Mushroom genome [Re: shroomed]
    #2152721 -

I'm down with the psilocybe mushroom genome project if anyone's willing to give a grant....

Noone has ever done this, to my knowledge, because it's too expensive and noone wants to support a project that will enable people to make psilocybin honey mushrooms, or psilocybin oyster mushrooms. It's not the main focus of "straight-minded" individuals.

If people with access to a lab were down, though, I could definately help....

--
Micro


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Any research paper or book for free
(Avatar is Maxxy, a character by Mizzyam, RIP)

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Re: Mushroom genome [Re: micro]
    #2152970 -

Thats good to know. :smile:

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Re: Mushroom genome [Re: Effed]
    #2153043 -

Is anybody actually serious about this? I really think it's possible, but it will take experiments, like western blots and stuff, and primers will have to be designed, etc.

This is probably the best question I've heard on the topic, because if one were to know the sequences for the enzymes on the Shikimic Acid Pathway after where Saccharomyces leaves off it would be really easy to make psychedelic beer.

The fungi that make beer would make psilocybin, too.

Ligating the DNA into the cell would be easy -- Saccharomyces electrocompetent and chemically competent cells are readily available -- the only task is to find the sequence of the enzymes on the metabolic pathway (which can be done with western blots, HPLC, etc.,) and then you are set....

--
Micro


--------------------
Any research paper or book for free
(Avatar is Maxxy, a character by Mizzyam, RIP)

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Re: Mushroom genome [Re: micro]
    #2153643 -

As far as mapping entire mushroom genomes, the most fully sequenced species are the model organisms Coprinopsis (= Coprinus) cinereus and Schizophyllum commune, but genome mapping is far from complete. Of course, these species aren't psychoactive. The only gene sequences for Psilocybe species on GenBank are boring old nu rDNA.

However:

"Attempted Molecular Cloning of Enzymes from the Psilocybin Biosynthesis Pathway in Psilocybe tampanensis"
a thesis available at:
http://meltingpot.fortunecity.com/gregory/1042/index.html

I stumbled across this some time ago, but as I haven't read it yet, I don't know if it has merit. I have a vague recollection of a paper I once saw, that characterized the psilocybin-producing metabolic pathway in some detail. Maybe there are some useful references in this thesis? I'll try to check it out after I post - it's a weird format.

If we have a situation where we don't even know what enzymes are involved in psilocybin production in the first place, then western blots / HLPC aren't of much use to us yet. A biochemist might suggest a few likely enzyme families, but let's assume no prior knowledge.

Obviously, psilocybin isn't a protein, so the issue is how you'd identify the genes encoding the enzymes which produce it. If I was starting from scratch, I'd collect mRNA trancripts whose abundance was correlated with levels of psilocybin production at different developmental stages of the growing fungus.

Then, you could immediately start playing around with transfections using DNA reverse transcriptions of these transcripts and hope for the best.

Check out this webpage on precisely this topic: Preliminary Feasibility Study for The Biological Production of L-Dopa, Mescaline and Tryptamines by Intact Recombinant Yeast Cells Using Only Common Amino Acids as Precursors to Bioenzymatic Synthesis
http://www.mescaline.com/misc/

Note the quote of $10,000 - 15,000

A more exacting approach would be to determine the endogenous loci of transcripts using appropriate probes, and then selectively and sequentially disable genes at these loci, while testing for continued activity. Visualizing loci would be quite difficult due to the unusually small size of mushroom chromosomes.

Some more potential problems: You might have a situation where the timing of enzyme release and supression is crucial to pathway function, in which case simply plugging in all the genes involved will not work, without their associated control factors. Also, interference or inconsistencies between the biochemical environment of the mushroom and the recipient may inhibit pathway function, even if the recipient is as closely related as a yeast. In a highly complex system you'd need to do some hardcore biochemistry. But I doubt any of these issues would be major obstacles in the case of psilocybin production.

In fact, as Micro implies, the whole thing seems temptingly feasible. All you'd need is one moderately wealthy shroomhead and one willing molecular biologist. They might even happen to be the same person.

But if I were to go down the route of genetic manipulation, I'd be interested not only in transgenic engineering, but also in endogenous over-expression factors. In other words, stick with growing psilocybes, but induce them to produce maximum levels of alkaloids, so that they literally exude pools of the stuff as they grow.

Suitable over-expression factors have already been developed by the Agaricus bisporus industry. You'd still need to know exactly where to stick them though.

Any interested benefactors out there?

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Re: Mushroom genome [Re: pluteus]
    #2154151 -

Quote:
A more exacting approach would be to determine the endogenous loci of transcripts using appropriate probes, and then selectively and sequentially disable genes at these loci, while testing for continued activity. Visualizing loci would be quite difficult due to the unusually small size of mushroom chromosomes.


I don't know much about this subject... maybe just few silly questions:
Is it possible to alter the exact locations or are they doing random changes and waiting until they hit the right one?
How the hell do they visualize loci?  :ooo:

Quote:
Any interested benefactors out there?


I'm going to buy a lottery ticket. :lol:

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Re: Mushroom genome [Re: pluteus]
    #2154164 -

I repacked the document. It?s now one big zip file.

Attempted Molecular Cloning of Enzymes from the Psilocybin Biosynthesis Pathway in Psilocybe tampanensis.zip


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Grow Mushrooms from Spores - PF-Tek for Simple Minds

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Re: Mushroom genome [Re: Anno]
    #2154359 -

Quote:
How the hell do they visualize loci?



I think pluteus is talking about comparative gene mapping. You can compare it with a like genus/species that's been mapped and add a tag there and then go up and down that part of the genome. You'd probably need a specific probe, though. When you run a southern blot on the DNA, though, the part with the probe lights up.

Pluteus -- very good post! (5 shrooms 4u)

Quote:
If we have a situation where we don't even know what enzymes are involved in psilocybin production in the first place, then western blots / HLPC aren't of much use to us yet.



Yeah -- I was thinking since tryptamine increases psilocybin production and decreases phosphorylation that's a good start already. I don't know how much more active the enzymes would be, though, and exactly how transcription is affected, so the bands might not be different enough, except the phosphorylase, of course, which should be almost non-existant in the 2nd set, as long as transcription is inhibited. Otherwise, simply inhibit trancription of the major enzymes like tryptophan decarboxylase or even chorismate or before (also present in Saccharomyces which is mapped,) which shouldn't interfere with any vital life functions. RNAi could be used to make this a lot easier. Then isolate the proteins and map them the best you can, then maybe compare them to a database?? Or maybe just sequence the DNA from the southern?? This would be a hit-or-miss situation, as my boss agrees, but given enough hit-or-miss situations, something is bound to work....

Sound at all feasible?

P.S. -- I'll read that paper after I take care of my cells.

--
Micro


--------------------
Any research paper or book for free
(Avatar is Maxxy, a character by Mizzyam, RIP)

Edited by micro (12/03/03 08:08 AM)

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Re: Mushroom genome [Re: micro]
    #2154470 -

Hey -- thanx for that paper! That's a LOT of good info!

I'll have to print it out so I can read it all, later. It's a really good paper, though -- thanks again for the link!

--
Micro


--------------------
Any research paper or book for free
(Avatar is Maxxy, a character by Mizzyam, RIP)

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Re: Mushroom genome [Re: zeronio]
    #2155569 -

Haven't found the wealthy shroomhead yet, but I have found a molecular biologist shroomhead with unlimited lab access (and she's cute too)! I'll keep y'all posted on any progress.

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Re: Mushroom genome [Re: micro]
    #2155594 -

If we can isolate the genes, the beer would be relatively easy. The genes could just be spliced into a plasmid and inserted into the yeast cells. Basically, once the genes were in the plasmid, anyone who could pass a high school chemistry class could do it. Psychoactive beer......I like the way you think!

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Re: Mushroom genome [Re: pluteus]
    #2155604 -

Thanks for the links, I'll check them out.

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Re: Mushroom genome [Re: rwpride]
    #2155991 -

micro - i'm glad the paper and post were helpful, unfortunately haven't had time to read it myself yet, your ideas sound very solid, too rushed for a more detailed consideration right now, but wouldn't it be great to collaborate on a project like this  :laugh:

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Re: Mushroom genome [Re: pluteus]
    #2167795 -

Psilocybin producing kudzu would be the project to get in on.

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Re: Mushroom genome [Re: Misnomer]
    #2168350 -

so where do you guys plan on starting? i mean what is step one priority action? and how can the community help? step two?

community needs a plan of what they should do if they are to help.

~JSlice~


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Re: Mushroom genome [Re: angryjslice]
    #2168438 -

Maybe try to reproduce that paper to see what went wrong? I'd have to read the whole thing, though -- I haven't had a chance, yet. If someone could do that we'd be set.

Money is the biggest problem, though -- that would require a good amount of money for custom stuff, so I kind of doubt it's going to happen....

Fun to think about, though. Maybe I'll do some experiments on my own, just to figure out some stuff about the enzymes....

--
Micro


--------------------
Any research paper or book for free
(Avatar is Maxxy, a character by Mizzyam, RIP)

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Re: Mushroom genome [Re: micro]
    #4237015 -

I amde a thread on this in advanced cultivation. I thinkt hatinstead of mapping the entire genome; it would be easier to produce a DNA sequence which would produce the DMT from amino acids. It would be rather easy. We already know/have the codes commercially available to produce tryptophan, decarboxilyze it, etc. All we really need to figure out is the dimethylation process. and the 4-hydroxyklation process.


The human genome has already been sequences, adn it is known that there are DMT-forming enzymes in the Lungs, blood, brain, and many other places int he human body. We could extract he knowledge from humans and put it in yeists.


4-hydroxylation is the tricky part.


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all posts made by 4hodmt are entirely ficticious. the user 4hodmt has noa ffiliation with any company, or organization; any endorsements made by 4hodmt are to be considered a joke. any similarity between a person (or story) 4hodmt is talking about is entirely happenstance. 4hodmt is not to be taken seriously under any circumstance. furthermore, he does not know what he is talking about. Please ignore anything 4hodmt has to say.


_______

"Trust me, I'm from the future.


Actually I'm not from the future, but I did lived there for a while."

- Baby Hitler

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Re: Mushroom genome [Re: 4hodmt]
    #4237049 -

I would certainly be interested in funding this project in the near future. as well as working on some stuff myself.

everyone interested int his should form a group so that we could discuss this further.


--------------------
all posts made by 4hodmt are entirely ficticious. the user 4hodmt has noa ffiliation with any company, or organization; any endorsements made by 4hodmt are to be considered a joke. any similarity between a person (or story) 4hodmt is talking about is entirely happenstance. 4hodmt is not to be taken seriously under any circumstance. furthermore, he does not know what he is talking about. Please ignore anything 4hodmt has to say.


_______

"Trust me, I'm from the future.


Actually I'm not from the future, but I did lived there for a while."

- Baby Hitler

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Re: Mushroom genome [Re: 4hodmt]
    #4237397 -

Interesting that this thread has been resurrected, I had forgotten all about it. It is now some years on and, thrillingly, we have a complete genome sequence for a mushroom species, namely Coprinus cinereus:

http://www.broad.mit.edu/annotation/fungi/coprinus_cinereus/background.html

The DNA sequence is largely unannotated and unassembled, but is already available in raw form on GenBank as accession number AACS00000000:

http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?db=nucleotide&val=33347384

I have done some initial work comparing some key Coprinus cinereus and Psilocybe genes that are popularly used in phylogenetics. Except for highly conserved regions (e.g. ribosomal subunits and spacers), there is a high amount of divergence in sequence composition and organization. This is to be expected, as there is no evidence to suggest that the psilocybe(/pychedelia) and coprinoid clades are particularly closely related. It will be fascinating to look for homologous genes in C. cinereus that are responsible for psilocybin synthesis in psilocybes. Of course, C. cinereus is not a psilocybin producing species, so if homologous genes are there, they will be dormant, defective, or involved in matabolic pathways having different end products.

None of this really helps 4hodmt's mission, which is anyway much more straightforward than identifying and transplanting the set of genes responsible for psilocybe alkaloid production.

I must point out that my interest in these matters is purely academic/hypothetical.

4hodmt, have you thought about the consequences of being the first person responsible for the unlicensed genetic engineering of a yeast to produce class A / schedule 1 hallucinogens? This is drug production on a level far beyond shroom cultivation.

You must be a very brave man. Count me out!

In any case I am personally uncomfortable with the idea of genetic modification purely for recreational purposes.

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