Gene Synthesis, Impromptu Discussion of Why Optimize Your Gene for Expression

Researcher shares frustrations of doing traditional cloning compared to using gene synthesis. Meeting room discussion covers pain points, comparison of costs, cloning multiple genes, complex cloning experiments, etc..

[William, Researcher, Synthetic Biology, Life Technologies] Leading in, we ask the question, "How many times has this happened to you?" You open the incubator only to find nothing but frustrating blue colonies. Repeated failed attempts to clone you know a large fragment puts you weeks behind in your research.

[George, Curious] How long does that process take? I mean is one attempt mean weeks behind?

[W.] So, if you do PCR and you might be able to do the cloning reaction in the same day and then you plate. Now day two I have a plate of colonies, I'm picking a colony, I'm doing mini prep to identify this is the fastest way to identify the positive transformant. I find it; I inoculate another culture end of day two. Start of day three; day three I will do my plasmid preparation. Day four I am sending my sample off for sequencing. So I would purpose that in the fastest scenario you are getting your experimental result in one week.

(...)

[G.] How often does that happen? [W.] For a large fragment, for a redo? Is what you're asking? I would say it's probably with our TOPO technology; it's probably about 50% of the time.

(...)

[W.] So gene synthesis is you type your sequence onto the computer and send it and if it's less than two kb you get 100% accurate, maybe optimized if you want it sequence in basically a week.

[G.] Why isn't everybody doing this?

[W.] So there's two factors, the first is that scientists are resistant to change. So even though this is an imperfect solution right now, it's what everybody else does, it's what I'm used to. I'm gonna stick with it. And then, there's the charge; the charge to do gene synthesis, the incremental expense over cloning.

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[W.] If everything goes right, and you strip labor costs out. The higher labor costs are associated manual cloning; when you push that into the equation, then they balance out basically. But the labor is always highly debatable.

[G.] What is the primary motivation for people to use gene synthesis, is that just the confidence that I'm gonna end up with a 50% chance that my results are going to be dead when I open the incubator?

[W.] Across the board the acts of the 100% sequence accuracy and basically freeing up my time to go do something else; that's the main crux.

[W.] So this has to do with the increasing need through systems biology for people to take multiple gene fragments and assemble them into a single construct. So the problem that happens with traditional experiments is that you end up with a crazy sequence of events to try to assemble one at a time each one of these pieces; that I need to get into my construct. So here we have basically a whiteboard with many different enzymes.

[W.] People still attempt it, they say "Okay, yeah, I'm gonna give this a shot. It's gonna take a week for me just to try and do the different assemblies, if everything goes right." So, we're saying end this sort of craziness of the complex experiments and then this is the gene optimization component. So you could do everything right and get your gene into the vector and then you put it into a cell and it doesn't express.

[G.] Is that with both techniques? [W.] No, this is with traditional cloning. [G.] Why isn't that the case with gene synthesis?

[W.] Because of gene optimization. So, this is the example of a human gene being expressed in a bacterial cell sometimes it just won't work. It could be toxic, there are a number of different reasons.

[W.] Then you find out it is not going to express. What if there was a way to get the exact sequence you need with 100% reliability, optimized to express in your host system in 5 working days? Would you be interested in that?

Now you can, with GeneArt Gene Synthesis and optimization, so then we're basically reiterating the points about gene synthesis gives you freedom from the burden of cloning, no more screening disappointing results. It's 100% accuracy every time. Confidence in your results, you don't need to sequence. You have the option to have your sequence cloned into your vector.

Our technology goes beyond just gene synthesis, goes into this realm of optimization so it's proven to increase protein expression levels. It does this is by stabilizing mRNAs , optimizing codon usage, and so on.

So we are now speed, no longer spend long hours waiting to get your construct. We have a SuperSPEED production option that can deliver 1200 base pairs in 5 days, 1800 in 7 days, and if you're in an emergency situation... so if there's a flu pandemic in the U.S... emergency genes available on request.

[mcpencil]

I struggle with efficient gene synthesis on a daily basis. Finally, someone has come up with a solution! Thank you, glorious LifeTechnologiesCorp!

[BlackHoleMan]

EPIC. I have been looking for a simple gene synthesis method.

[TheKylestanger]

this happens to me every day! thank you so much!

[doombybbr]

do many at once: large sample size, get a few people to help to speed up research.