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Molecular Cloning and the Synthesis of Recreational Drugs

jspun

jspun

Bluelighter
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Jun 11, 2008
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For my 50th post I choose a topic I've been interested in and curious about.

How feasible would it be to use molecular cloning techniques to engineer eucaryotic or procaryotic cells to develop a renewable source of recreational chemicals? In other words synthesize or isolate genes involved in the biosynthesis of a drug in nature, insert them into a vector, transfect into target cells, culture the transfected cells, then isolate the substance of interst. The genetic sequences for drugs like morphine, cocaine, DMT, and precursors like ephedrine, safrole, ect...are already out there in nature. What are some of the practical limitations involved in pulling this off? This is probably science fiction but think of how cool it would be to get ahold of one of these cultures.
 
This is has been discussed tons. Google will give you tons of information about it!

It's promising, but hasn't really been done yet.
 
Its a great idea.

I think it could be done with the right lab, all the tech and know-how already exists, it basically needs someone to do it. If you could just get a psilocin-gene-transfected acidophilus strain for yogurt, you'd be my hero.
 
This is has been discussed tons. Google will give you tons of information about it!
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tried googling it. Havent been able to find anything. What do you recomend I type in? Any links or threads on bluelight. Thanks!

If you could just get a psilocin-gene-transfected acidophilus strain for yogurt, you'd be my hero.
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That would be cool as hell!

A comon application of molecular cloning is to get bacteria to express luciferase. I think that this technology is used to produce antibiotics too. Maybe getting unicellular organisms like bacteria and yeast to produce something that is made by multicellular organisms could conceivably be more problematic from my standpoint ( with my limited understanding of mol bio).
Stuff like protein folding problems, getting promoters to work reliably, problems with accessory pathways, getting the biosynthetic pathways to work the way they are supposed to in cell based systems, and so on. You would have to feed your culture the proper growth media too.
 
why yogurt?

If you want a lot of something you start the fastest.

vectors seem like the best way to go about it.
 
Why yogurt?
1) Its easy to make yourself if you have the right cultures.
2) If made right, it tastes good.
3) It would be easy to load up a bunch of snacks to take anywhere and not get busted.
4) You could sell homemade yogurt cups on the cheap because the acidophilus would do the work of a fungal mycelium in 1/100th the time.

I could go on...
 
One would not be able to regulate the dose of course with tryptamine producing yoghurts, not effectively at least.

Electroporation might be a viable and practical alternative, it uses electrical pulses to temporarily open DNA-permeable pores in the cell walls, I'm looking into the details myself actually, if one could buy the plasmids for each gene nescessary (or otherwise isolate the genes) from a gene bank thn it should be doable with a lot more ease than if one were to have to DIY a genetically engineered phage (and safer)

Then should be as simple as zapping the correct elctrical voltage/current/frequency for a short time through a suspension of the DNA to be inserted and the cell culture in a suitable culture medium, allowing to recover, then spotting onto culture medium.

Beats buggering about with monkey kidneys and the like.
 
e coli is the standard for these things I use it all the time to express O-AcetylSerine(thiol)Lyase. Which is a plant enzyme, so its really not cutting edge, rather a common tool in MOlecular Biology
 
Electroporation is a super common technique. That is not the shortcoming. The problems are multiple-getting the proper enzyme's genes (we have no idea what most of the gene sequences are for most drug pathways, morphine being the partial exception), making sure they work in a heterologous expression system, regulating their activity so there are not bottlenecks, reproduce-ability for consistent yields, and probably some more I can't think of. It really is non-trivial even if we get the proper enzyme sequences in a few years because you have to make sure all of the pieces work together in an environment in which they were not evolved to (things like folding, post-translational modifications, allosteric modulation) and in harmony so as not to make too much or little of something and poison the organism.

I would recommend looking through the work of Jay Keasling @ UCB for more info including inserting and optimizing the artimisinin pathway into yeast. The papers make it look non-trivial but it is a paper (read nothing but success) and he has an ARMY of the best students and postdocs around.
 
Perhaps there's no one to fund it? Who would fund such research. I'm not a molecular biologist myself, but I know the department of mol. bio. at my university spends A LOT on the research they do. Thus I think its quite expensive developing such a method. Sounds interesting though :)
 
What about just makin' 'em make the enzymes, mass produced, using commercial plasmids containing ready-made insertion and transcription sites, as well as antibiotic resistance sites? Cost is an issue, but it makes it a ton easier to select for organisms containing the genes in question. I'd just chop the bacteria up after maintaining an active culture and run each stage by myself. Problem is finding the genes that code for the enzymes needed. Do we even know what enzymes in, say, mushrooms, do the job, let alone where the genes are? These aren't sequenced organisms yet.
 
No disrespect intended- but this is perhaps the WORST way to produce drugs. I did my PhD in producing novel proteins in E coli and, believe me, you don't wanna go this route to produce anything. First off- you'd have to isolate the drugs from the millions of cellular proteins and other shit present upon lysis of cells. Also, E coli and eukaryotic systems are only of any value to produce proteins that are fairly big (ie. ones that can't be synthesized). Scaling up cellular systems for production is also REALLY REALLY HARD!!!! I understand where you're coming from- but this just aint a good idea at all. NEVER deal with cellular systems when a synthetic route is possible. That is ALWAYS THE WAY TO GO!!! The headaches man...the headaches... no way.
 
^ Believe me, I totally get what you are seeing. But, what of all the work people have done in the past to create short cuts!! Dont you think some guys said 150 years ago "What the fuck are you spending so much time making explosives move you around, when a horse can do it right now!!"

I think the biggest hurdels have been said. Finding all the genes that code for the enzymes that make the drugs, and then like you said mass production, then isolation. But dammnit, give me the shit and Ill HPLC that to death and give you pure product!


But I think 5 years of research can end up mapping the production pathway from drug to gene in a plant
 
Of course the ultimate dream would be to use a heterologous expression system with various proteins from Claviceps p., reverse-engineer the reaction-mechanism and molecular dynamics of those enzymes and then, using site-directed mutagenesis, develop a non-fungus in vitro biosystem that directly produces LSD with no other ergoloid side-products (and no ergopeptines). Hell, even something that produced ergometrine or hydroxyethyl-lysergamide (LAOH) would be adequate for a nice renewable source of psychedelic ergoloids.
 
theseeker said:
No disrespect intended- but this is perhaps the WORST way to produce drugs. I did my PhD in producing novel proteins in E coli and, believe me, you don't wanna go this route to produce anything. First off- ...etc...
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There's a significant difference in producing proteins vs. non-proteinic small molecules via a biotechnological way, the latter one usually being easier. Comparing apples with oranges, I'm afraid...

- Murphy
 
Yeah. If you want to produce proteins you're not going to have a hard time at all.

Novel proteins, though, is not the same either. By novel, you means proteins that hadn't been seen before or proteins that weren't produced naturally before?

Protein synthesis is getting a lot better, so this probably isn't a real bright way to go either, for those, except for substantial sized things. You wouldn't bother with endorphin-producing E coli.

I'd like to see amfonellic acid producing e coli though!
 
Everyone has interesting points. But I really don't see the added value of doing such a synthesis in cells. As a poster above pointed out, it would require the synthesis of multiple enzymes (I don't know the pathways however). In E coli, only about 30% of heterlogous proteins introduced by plasmid vectors can be expressed. It isn't even always possible to express heterologous proteins that E coli produces itself!! Yeilds are typically low- even from T7 driven transcription. Also, some proteins are better (or can only) be expressed as fusion proteins.

Now, if you want to tell me that you'd like to produce enzymes that will limit side-products during the synthesis of drugs- I can understand that. Even then, though, yeilds will be low and the activity of the enzymes would be in question.

Interesting topic.
 
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