[azureus]

We're a nanopore sequencing shop. Couldn't agree with you more that nanopore generally is the future, not just for sequencing. Can't wait for ONT's solid state nanopore flow cells - you may get flowcells that go 2/3 times as long then.

But then for most things .. the problem is the prep, not just because the whole portability thing goes for a toss. Yay, great the sequencer is the size of a USB drive, but the rest of the lab isn't :/

More worryingly the biggest and most stubborn cost is now the prep, not the sequencing.

As you correctly pointed out - need to find a way around amplification - then both of these problems above go away if you can do direct PCR free sequencing.

The other less mentioned problem related to the above is also the need for parallelisation - some of those ultra low costs you read about can only be realised when you sufficiently multiplex your samples. For instance, its about 100 USD per reaction for ligation (last step of prep before the sequencing starts), you generally wait till you're sequencing atleast 12 samples in the same reaction so that you're paying <10$ per sample, not 100$ per sample, which is obviously insane.

[koeng]

The key is to do barcoding at the PCR amplification step. That way, you can get away with barcoding hundreds of samples in a single tube.

Really the prep screws it any other way. Is ONT coming out with solid state? Got a reference?

[azureus]

Solid state nanopores: still in research: https://nanoporetech.com/how-it-works/types-of-nanopores

I'm pretty sure they will come out with it. The protein nanopores were the first wave of nanopore research - its tried tested and stable now so they stick with it.

A few years after they launched, the first solid state nanopores were being demonstrated in the lab. Commercialising solid state nanopores seems to be easier if anything than protein nanopores because they slot right into silicon fabrication.

On PCR/barcoding .. Yeah, thats right - do it right in the PCR step .. sometimes we avoid it, if we are not yet sure about the protocol. I think what I meant to say is that the full promise of nanopore sequencing for me is achieved only when you can skip having to amplify/multiplex/barcode - just extract dna, wash, add sequencing adaptors and go - for almost anything.

I think the way they are talked about people generally come in expecting that TODAY .. they think they can literally stick a single sample with no prep and get 1gb of sequencing done for 10$ in an hour. I've seen quite a lot of that. (even from people with PhD's :) )

So yeah its more that the minute you go PCR, you're in for a minimum 20$ per sample, often its the highest cost line item in your whole process.

If you're doing things like 16S metagenomics, you get sequencing at 2-3$ and prep at 10 times that, starts to feel "wrong" after a while, if you get what I mean.

Why're trying everything we can to make sure we're running at full capacity so that we can always give low prices even if its single samples.

Also seeing if we can reduce prep cost with microfluidics/mems - they have voltrax for this, but there are a few other vendors in the market. That has the positive knock-on effect of also reducing labour cost.