Clive G. Brown
@The__Taybor
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Excellent advice.
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We use a lot of our own flow cells. So we have custom vending machines to store and dispense on demand.
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it official, MinION is the worlds best selling NGS sequencer (now outstripping MiSEQ).
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not PCR-ing your genome, gives near flat coverage on Nanopore (unsurprising as we randomly sample long frags from soln). Every sequencer type that depends on amplification suffers GC/AT PCR biases in coverage, and lower coverage regions have the less confident mutation calls.
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GitHub - nanoporetech/taiyaki: Training models for basecalling Oxford Nanopore reads - now including modified bases.
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Progressive 2 step push into reaction column. Extraction, then bead drop into flowcell. ~5-10 mins.
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Nice graphic, right on the lower end there you could put a single nanopore, ~10 nm
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just stumbled across a slide form Q1 2011 Board meeting where I floated MinION concept ...
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I know its just a graph. But we can now get 200G out of a PromethION flow cell. (Thats 2x 30fold WGS at ~$350 each).
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I am hoping and optimistic we can launch our single cell sequencing device in 2018.
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A late xmas.
3 runs of PromethION cells, two in progress. One at 90G in 65hrs, another on track for 90 in 45 hrs.
thats equiv to 30X human for < $1000
~$680 cell + ~$70 sample costs. No capital.
90 * 48 cells = 4.3 Terabases (> NovaSeq)
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We've had 62G off a PromethION cell, i'd like to now see 90G for my xmas pressie.
that would be a 30X human genome equiv. on a ~$600 flow cell.
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GitHub - nanoporetech/taiyaki: Training models for basecalling Oxford Nanopore reads
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Previous PromethION cells in blue, new ones in red in mid run with active unblock - almost 90G in 24 hrs
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Preliminary R10 on promethion, tweaked chem and callers but running slower ~ 300 bps. Q20+ mostly, we think we can kill the tail.
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We have more than doubled duplex rates. More to come from tweaks and software (~80%) . A non disruptive roll out is possible.
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70+ pores (I think more) per well. 96 wells. Custom plate with embedded liquid biopsy type sample preps and enrichment. Test/screening oriented.
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There’s a 1 Megabase Q30 in read in there. If one can be like that they can all be like that. That must be the longest and most accurate fragment of DNA ever sequenced.
What’s more, telomere-to-telomere (
#t2t
) assemblies now achievable with JUST simplex.
Q28 simplex data is accurate enough.
You do not need data from any other platform — paving the way for
@nanopore
T2T assembly, using just simplex data.
#nanoporeconf
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Whats this? Some early R&D data. Now getting around Q20+ at 415bps. Yes faster chem and Q20+… more training/tuning required.
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Adaptive-sampling ['Read-Until'] coming mainstream in the next MinKNOW. Any panel - any organism.
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We’ve kicked off a new project at ONT. ill be transparent given what’s going on, interested in feedback (dont want to waste time). 1/n
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Some recent P48 runs. GIAB human.
7.3+ Terabases.
Still <= 50% of what’s possible at 450bps.
All 48 cells running, live base caller.
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Latest Guppy went out yestderday, about +2% raw-read accuracy increase, about +Q5 on consensus R9.4, fast to run.
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Recover library from Nanopore flow cell whether successful or a fail. Re-run, re-process, don't lose a precious sample.
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Adaptive Sampling [read until] now fully released in MinKNOW
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Proto board for USB base calling accelerated FPGA dongle. Will shrink.
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Back of Envelope; 1 PromethION can do 100k COVID genomes per 24 hrs at around $7-$10 per sample. Thats without Flowcell re-use. Pipelined sample prep [pcr] needed at speed, probably limiting. Assume 96 native barcodes.
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base calling and RNN training tools, nascent consensus calling, have been open sourced, as-is, no warranties, wip.
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Some base-caller updates coming within 5-10 days. 98%+modal and many reads above Q20.
Note the X-axis.
Generally, sig +ve uplift in consensus and mutation detection.
Slightly slower speed in research version.
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GPU accelerated "Guppy" base caller now on poretal - Developer only for now.
~800K bases per second / GPU
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GitHub - nanoporetech/megalodon
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Amgen invests £50 million ($66 million) in Oxford Nanopore Technologies
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And .. the latest series-D asic MinION/GridION flow cells.. up to 30G … note the extended runtimes.
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Here's m'draft genome 'selfie', at least a white blood cell branch. All MinION.
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'field kit' - lyophilised reagents for use away from lab, live tomorrow. no cold chain.
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There we go, long predicted ;). Begining of the end of short reads. ONT not on the menu for them, who else would they go for. Suggests internal long read tech innovation not imminent.
Suprise, genomics folks! $ILMN is buying $PACB for $1.2 billion. I spoke to CEO Francis deSouza about why.
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Latest from Akeson al ; Reading canonical and modified nucleobases in 16S ribosomal RNA using nanopore native RNA sequencing
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Our latest MinION runs. 16G now, just a MinKNOW upgrade away.
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I had some fun this week trying to convince somebody that Nanopore is actually the most accurate DNA sequencing chemistry ever developed ....... on a per molecule basis.
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People who are good in the lab are usually good in the kitchen.
Cooking classes give science students a taste of real world
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Tims first cut of the RLE base caller we mentioned at LC.(GPU version to follow)
GitHub - nanoporetech/flappie: Flip-flop basecaller for Oxford Nanopore reads
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Compression for nanopore signal.
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Anyhoo. There’s some recent MinION runs at 40G. Team think we’ll smash 50 soon. Release follows.
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Each point is a new pore-motor combo. Y=acc, X=speed. The graphs below are the hatched green, some optimisation applied, the others no so much.
Blue is variants on what you have.
Whats this? Some early R&D data. Now getting around Q20+ at 415bps. Yes faster chem and Q20+… more training/tuning required.
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Over 30G from a MinION flow cell in the field. Thats news.
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Further, 7.1G in 30 Hrs on next releases, should get ~10G in 48 hrs with some scripting tweaks. Tech update soon.
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somebody tell him ....
Today we're sharing research from
@StanfordMed
on how the genome sequencing method we built with
@ucscgenomics
helped speed up identification of disease-causing variants, enabling faster NICU diagnoses.
#GoogleAI
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Started shipping the ‘full-on’ PromethION 24s and 48s. Quiet, all flowcell positions enabled, onboard analysis.
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Highly parallel direct RNA sequencing on an array of nanopores | bioRxiv
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Single port flow-cells out today for WGS junkies. I think slightly more channels also ...
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I think its out :
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Illumina and Pacific Biosciences Announce Termination of Merger Agreement | Seeking Alpha
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Adaptive sampling [‘read until’] in a simple form for everyone via MinKNOW. Panels, subsets, regions etc etc.
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VolTRAX V2 is just starting to ship. Thanks for patience, ramping up over the next couple of months.
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Blimey. we're in the Daily Mail
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Reminds off a visit to a genome centre some years ago, they asked: ‘aha! But you can’t do read-pairs can you!’?!. I said ‘ yes, easy, ill take a 49kb nanopore read, and delete 48.6kb from the middle of it and leave you the ends’.
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I realize this might get boring but latest PromethION cells at 90G (30X human equivalent) - in 36 hours, down from 48.
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‘Field Kit’ launched by Oxford Nanopore, enabling shipping and storage of sequencing preparation kits without reliance on cold chain
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UMI type high accuracy multi-pass reads now supported by ONT.
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Firstly, and probably of general interest to existing customers, we have been working on improved base-caller models, training sets and a tweaked motor. On SIMPLEX, natural DNA, at 260bps we are seeing the distribution as shown. This is subject to more validation. Yes, thats
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MinKNOW 1.10 released. Raw RNA calling. Writing to network drives.
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A favourite of mine. Motor is fused to pore, catches natural DNA from soln. Possible to remove many upstream steps, maybe all.
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@StevenHailAus
@vporras95
The sun, fusion, drives most energy capture on the surface and that is effectively infinite- then you have nuclear fission and decay to tap into from terrestrial sources. With abundant cheap energy you can recycle - your assumptions also based on closed system of earth - mining
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and there it is
#cliveome
- all three rows on this Beta. (24 x 150G is 3.6 Terabases) less 4 long frag preps.
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Release v0.5.0 · nanoporetech/bonito · GitHub
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A MinKNOW update is out today which improves early yields in runs.
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Planning the next generation ASIC formats today. Easiest thing is 10X number channels/pores per unit area, all products, but does that make sense ? [assume 800bps] - e.g. 5Tb per PromethION cell, 600G MinION etc .. ? Alternative is new formats.
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Tim has updated the walkthrough for base caller training :
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a 24 cell PromethION run (Good sample) - but only 22 Hrs. Still 1.3 Terabases. Expect over 3Tb in 50hrs. Run is on.
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direct RNA kit was launched today, but also note cDNA system relaunched with performance like DNA (i.e. millions of reads full len tscpts)
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Nanonet goes open.
Example RNN implementation (but not definitive). Read simulator to follow for those without data
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#cliveome
2.0 (or is it 3.0) is kicking off today. On PromethION. Aiming for 2-3Terabases - at least 1 sub $1000 flow cell at 30 fold. Mix in some ultra longs. Data will be public. Might do some 1D^2 (revamped).
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We have cleared the Flongle back log, now producing more than the order rate of premium grade flowcells. Today we discussed bringing the order to delivery times right down.
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OK these things are boxed up and ready to ship in numbers ....
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field sequencing kit (rapid) announced :
broadly avail soon.
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Flappie is coming to developers imminently, replacing Scrappie. R9 versions first, then followed by R10 versions within a week to two (ish). R10 itself is now under field test. Updated consensus callers also coming out in sync. Flappie can optionally support direct mC calling.
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Some more promethion cells, 120G. Sharp eyes note the ultra run times.
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Cliveome with new long read method. Hope to release protocol soon. New normal ?
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Dorado will replace Guppy.
Happy to share our first release of dorado 🎉
- Multi GPU support
- Latest quantile scaling
- Automatic parameter selection for best performance
Plus a whole lot more!
Make sure to try it with Pod5 to really see it fly 🔥
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