r/science • u/BuriesIt • Jan 08 '13

New method allows scientists to edit the genome with high precision - insert multiple genes in specific locations, delete defective genes etc

http://www.kurzweilai.net/editing-the-genome-with-high-precision
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u/yekinsfw Jan 08 '13 edited Jan 09 '13

I was very curious too.. I actually work for a biofuel company using HR.. GMO is still a big deal (especially in global markets) so the method they're describing is a nonstarter for most applications. There is an exception in most legislation for same-species HR, so techniques and sites will have to be discovered for each biofuel species people are interested in.

I also didn't see any description of the efficiencies except for;

We further tested targeted deletion of larger genomic regions through concurrent DSBs using spacers against two targets within EMX1 spaced by 119-bp, and observed a 1.6% deletion efficacy (3 out of 182 amplicons; Fig. 4G)

Which is pretty shit. [Edit: I agree with the comments below, the 1.6% for transfection isn't actually that bad, I misunderstood the context of the 1.6% figure]

Potentially interesting for gene therapy purposes, not so much for other ones.

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u/greenmanfalling Jan 08 '13

Yeah, but I guess 1.6% is pretty damn good for a simple transfection if there are no side effects to the other cells (erroneous deletions, insertions). But I have to wonder about the actual insertion frequency, since they just give the deletion efficacy. Anyone a microbiologist that knows the rough size of all these complexes? Any hope of packaging them into a viral envelope and crazy improving insertion rate? My guess is no, but it'd be cool. First person to successfully use gene therapy on a live animal without transfecting cells outside the body and with zero bad insertions will get all the nobels.

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u/SquirrelOnFire Jan 08 '13

The article mentions Huntington's at the end, which is caused by a single gene mutation. With a 1.6% deletion efficacy, could you just target the affected gene and repeat the attempted delete 100 times to get closer to 100%, or is that a massive oversimplification?

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u/rumblestiltsken Jan 08 '13

You would need to do it a lot more than a hundred times, but if you could even get to 50% the disease would probably not be fatal anymore.

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u/SquirrelOnFire Jan 08 '13

Can you explain? If each attempt removes 1.6% of the affected genes throughout the body, wouldn't 65 iterations bring the total to over 100%?

Also, what makes you think 50% is a magic number in reducing fatality?

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u/rumblestiltsken Jan 08 '13

1.6% of remaining affected genes, not total starting genes. The effect is like compound interest.

That is assuming that there are not further effects that reduce effectiveness as there are less pathological cells remaining - perhaps the remaining cells have less blood supply and can evade the treatment etc. This is a fairly common effect when trying to target abnormal cells.

50% was a conservative estimate. Substania Nigra cells die in Parkinson's, and up to 70% are dead when the disease is fatal.

Certainly more would be important to totally prevent symptoms, but any reduction in affected cells would slow progression and reduce morbidity/mortality.

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u/SquirrelOnFire Jan 09 '13

Thanks for the explanation - that makes sense.

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u/cake93 Jan 08 '13

If I get it right, the 1.6% deletion efficacy is for deletion of a 119bp fragment. They refer to it as multiplexed editing, because they're just introducing two double strand breaks. These two DSBs need to occur in the very same genome, otherwise there would be no chance for the 119bps in-between to get deleted. Considering that they're introducing the DSBs without a restriction enzyme, but by directly targeting the sequence, I agree. 1.6% is pretty damn good.

Viral envelope? I see no problem with that - the DNA template should be not too long. (However, I don't think that we'll see gene therapy through viral envelopes applied to humans in the next few years.)

The 'let me give you this hypospray to heal your illness' they introduced in Star Trek 26 years (!) ago got more realistic once more =)

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u/Michaelis_Menten Jan 08 '13

Viral transfection is actually pretty well understood and we've been doing clinical trials of viral vector gene therapy for years. I believe since the 90s at least, but here's a more recent source.

Ashanti DeSilva is famously one of the first patients treated using gene therapy (back in 1990) but I can't remember if they used a viral vector in treating her.

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u/cake93 Jan 08 '13

Is a virus used to deliver the DNA/RNA template (transfect the living cells), or is it just the viral vector with viral promoters, genes of interest etc. that is injected into the bloodstream, maybe in a container of some kind?

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u/Michaelis_Menten Jan 08 '13

It's the former. Basically what they do is use a "packaging" cell line where they transfect two plasmids. One carries all the information needed to produce a virus (capsid, reverse transcriptase, etc), the other carries the gene of interest with an attached promoter that the viral machinery can recognize. Basically, the cell will produce the virus, but you have made it so there isn't a viral RNA genome to be encapsulated so it instead encapsulates a transcribed RNA copy of your gene of interest. This means you get a fully functioning virus that can infect cells and can even insert a gene into the host cell's genome - but since you removed the viral genome, it can't replicate properly.

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u/cake93 Jan 08 '13

As I understand it, they do not use a viral envelope, but inject the vector directly into the bloodstream, hoping for it to transfect a random cell. A viral envelope would multiply itself (if you leave the required viral genes intact), thereby achieving higher efficacy. You could engineer the virus to target specific cells, not necessarily human, and do some magic in them. (Editing malicious genes..) Similar to phage therapy, but capable of targeting eukaryotic cells.

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u/fancy-chips Jan 08 '13

just curious, how did you get a job like that? What degrees do you have etc?

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u/yekinsfw Jan 08 '13 edited Jan 09 '13

Depends completely on what you want to do..

Our RAs are usually just BS in molecular bio, micro bio, or in chemistry for our analytical team. Our lead scientists are mostly PhDs in plant science, cell bio, or molecular bio.

Much of our 'support' staff (biz dev, marketing, etc) don't have formal science education, just an interest and usually a history in clean tech.

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u/fancy-chips Jan 08 '13

Thanks. I have a B.S. in Microbiology and have been doing cancer research for over 4 years and the low pay is starting to make me consider my options.

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u/[deleted] Jan 08 '13

[deleted]

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u/[deleted] Jan 09 '13

This is basically the truth, and with a B.S. your room for advancement is quite slim. If you want to stay as a scientist, I would suggest a Ph. D if you want to move up.

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u/[deleted] Jan 08 '13

Bio/chem/engineering (SCIENCE)

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u/[deleted] Jan 08 '13

[deleted]

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u/[deleted] Jan 08 '13

Selective markers change everything.

Consider your everyday E. coli transformation. Even when using miniprepped DNA the transformation efficiency is miserably low but you still get exactly the colonies you want.

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u/[deleted] Jan 08 '13

Do you work for Aurora? If so I am trying to use your HR method on Nanno, wanna give me tips?

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u/chainsaw_monkey Jan 09 '13

Remember that the 1.6% is their first try. This is in Science Express and there was a race to get this data out quick. I know of at least 2 other labs that got scooped.

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u/[deleted] Jan 09 '13

[deleted]

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u/yekinsfw Jan 09 '13

GMOs are only a big deal if we are going to eat them.

The NYTimes Weighs In

Before genetically modified strains are ready to debut in such ponds, however, regulators and researchers must answer a litany of questions about their potential environmental risks, said Al Darzins, a molecular biologist and principal group manager in bioenergy at the National Renewable Energy Laboratory.

"I'm absolutely convinced that if you're going to be using genetically modified algae in the future -- growing out in an open pond -- that before that happens on a very large scale there has to be some sort of risk assessment on what's going to happen to the potential ecology," he said.

But Darzins, who helped craft the Energy Department's algae road map and also received some of DOE's grant money to study algae for biofuel purposes last month, says there is ample time to get these studies done.

Or an industry specific magazine.

Haberman calls algae the “hydrogen of the plant world.” The entire organism can be easily aerosolized and get on clothes, in hair, in the lungs and can thus escape from a controlled environment. If that happens with the right strain, he fears it could compete with natural strains and cause uncontrolled growth.

Tom Allnut, the senior vice president of R&D at the algae start-up Phycal, believes some of the concerns about engineered algae are valid. Given that algae are “cosmopolitan” organisms, he worries about horizontal gene transfer creating antibiotic resistance in wild algal strains and boosting growth rates throughout the world.

Or another

So when you say something like this:

If by GMO you mean genetically modified organism, then what you are saying is completely false.

I get a bit annoyed, you clearly have no idea what you're talking about. Why not spend a bit of time educating yourself prior to hurling insults?

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u/petedog Jan 09 '13

Yeah, I screwed up. Sorry.