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Oxford Nanopore MinION – USB stick-sized DNA sequencer

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Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#11
post #5

Earlier quoted context omitted.

No, it does not. You still need a wetlab to do sample preparation. There are a couple reasons why this is exciting though. Oxford Nanopore has seemed like bogus vaporware for several years now, and many people were skeptical that they had a viable product. This data release is the strongest suggestion yet that the thing actually works (but is still in no way conclusive). Two, assuming it does work, being able to sequ…

How portable is a "wetlab"? Could I take this with me into the jungles of south america or the se asia?

A wet lab is a lab as you might imagine it on CSI, i.e. the whole nine yards.

Sometimes, components of this can be miniaturized to create a "lab on a chip", but that takes another level of technological development.

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#12
I would like to see what the real raw data looks like. There doesn't look to be any noise in that wiggle plot and the signal has already been processed to a degree (see the hard edges and squareness of the features). That's really the hard part though, going from raw to digital, and the decisions that are made regarding how that transformation is achieved will have a lot to do with the quality of the data.

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#13
post #8

The error rate in the "good part" is about 15%. That is quite frankly very very very poor. This might be a good alternative to pacbio, (unless the other technologies can increase their read lengths). Typically (I chatted with my contacts at the Venter Institute)... best results are gotten by making long reads using pacbio or (maybe minion, they are looking into that currently) and using that to generate the scaffold…

More information: For personal (as in human genome) sequencing, I'm not entirely sure how useful this is... We already have a scaffold for the human genome (that would be Mr. Venter himself) and MAYBE you could get some haploid information out of it, but you'd worry that the base pair you care about for any given individual is in one of the wrong stretches. For organism sequencing, I think this could be even worse th…

Naive idiot alert.. could you process the "same" DNA numerous times then take the mode in each case? Or are the errors essentially ones that would be repeated each time?

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#14

Earlier quoted context omitted.

More information: For personal (as in human genome) sequencing, I'm not entirely sure how useful this is... We already have a scaffold for the human genome (that would be Mr. Venter himself) and MAYBE you could get some haploid information out of it, but you'd worry that the base pair you care about for any given individual is in one of the wrong stretches. For organism sequencing, I think this could be even worse th…

Naive idiot alert.. could you process the "same" DNA numerous times then take the mode in each case? Or are the errors essentially ones that would be repeated each time?

[deleted]

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#15
post #2

What does it mean? I don't suppose I can put a drop of blood onto that sequencer and receive a sequence?

No, it does not. You still need a wetlab to do sample preparation. There are a couple reasons why this is exciting though. Oxford Nanopore has seemed like bogus vaporware for several years now, and many people were skeptical that they had a viable product. This data release is the strongest suggestion yet that the thing actually works (but is still in no way conclusive). Two, assuming it does work, being able to sequ…

[deleted]

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#16
The error rate of 15% (I actually believe it's more like 30% in practice, which is 2x PacBio's) is actually excellent and comparable to PacBio's in that the error is primarily randomly distributed, and not systematic like current Illumina or IonTorrent systems (which have admittedly lower error rates, at 1-3%). The short of it is, absolute error rate is not the whole story.

When looking at what ends up being just a massive string of 4 characters, having 15% error on a particular read doesn't matter too much; you end up getting many reads (fragments of a full sequence) which overlap the same area of the genome. With enough reads (and having many, say 15-100x coverage as it's called, is not uncommon practice in sequencing), this error is obviated by consensus. The long read length is especially useful for this, as the lengths which overlap are much greater than with current NSG reads of 150-200bps.

Additionally, I think the goal of the MinION were more to display a sort of disposable (sub-$500) machine which could be used on-site and without sample-prep. A forensic kit, really (though probably more for creatures such as bacteria and fungus rather than humans—I recall that it would take 2-5 MinIONs to sequence a human genome to even a short depth, but I may be misremembering).

In short, this is a cool proof-of-concept for Oxford Nanopore, as their technology has been talked about for a long time with nothing to show until now. It will be interesting to see where it goes next, particularly if the reads lengths keep climbing and the error profile drops even more.

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#17

Earlier quoted context omitted.

More information: For personal (as in human genome) sequencing, I'm not entirely sure how useful this is... We already have a scaffold for the human genome (that would be Mr. Venter himself) and MAYBE you could get some haploid information out of it, but you'd worry that the base pair you care about for any given individual is in one of the wrong stretches. For organism sequencing, I think this could be even worse th…

Naive idiot alert.. could you process the "same" DNA numerous times then take the mode in each case? Or are the errors essentially ones that would be repeated each time?

Actually PacBio does just that to get better a better error rate. They basically have X amount of sequencing that can be done. You can spend that X however you'd like. If you want a sequence that is X long, you'll have a higher error rate. If you want a chunk that is X/10 long, you can circularize it, and thus sequence it ten times. This gets you better accuracy with the redundancy. DNA is pretty robust.

In practice though, even with these "circular consensus sequencing" reads, the error model is significantly higher than other technologies.

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#18
post #16

The error rate of 15% (I actually believe it's more like 30% in practice, which is 2x PacBio's) is actually excellent and comparable to PacBio's in that the error is primarily randomly distributed, and not systematic like current Illumina or IonTorrent systems (which have admittedly lower error rates, at 1-3%). The short of it is, absolute error rate is not the whole story. When looking at what ends up being just a m…

As it stands, it looks good enough for identification purposes (for example of a pathogen), but nowhere near good enough for genetic studies where single base pair variations are the most sought-after aspect.

For human genetic studies, I guess it also depends on how readily multiple reads can be obtained, and whether the errors are randomly distributed.

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#19
post #6

Very nice! I worked on the firmware for the "Applied Biosystems ABI Prism 310 DNA Sequencer". I remembered it cost >$60K each when it was released ~1996, 1997 time frame. It is good to see DNA sequencer tech progress faster than Semiconductor.

> I worked on the firmware for the "Applied Biosystems ABI Prism 310 DNA Sequencer".

Do you have copies of that firmware or know where I could pick it up? I'd like to poke around, try some reverse engineering, etc.

Re: Oxford Nanopore MinION – USB stick-sized DNA sequencer

#20
post #17

Earlier quoted context omitted.

Naive idiot alert.. could you process the "same" DNA numerous times then take the mode in each case? Or are the errors essentially ones that would be repeated each time?

Actually PacBio does just that to get better a better error rate. They basically have X amount of sequencing that can be done. You can spend that X however you'd like. If you want a sequence that is X long, you'll have a higher error rate. If you want a chunk that is X/10 long, you can circularize it, and thus sequence it ten times. This gets you better accuracy with the redundancy. DNA is pretty robust. In practice…

I don't think they push CCS much anymore. The focus seems to be on generating long reads and then using Quiver to call the consensus (https://github.com/PacificBiosciences/GenomicConsensus/blob/...).
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