Oxford Nanopore is the first company to commercially launch a nanopore based DNA sequencer. Nanopores are small holes through which DNA can travel, generating an electronic signal that is used to sequence the individual bases. Oxford Nanopore sequencers, which are inexpensive to own and operate, are able to generate very long reads, albeit with a relatively high error rate. The long reads make them a good fit for applications like de novo assembly while their portability has made them the only practical choice for sequencing directly in the field.
The MinION is the first commercially available nanopore-based DNA sequencer. It is also currently the only true “handheld” sequencer, measuring 10 cm long and weighing under 100 g.
The PromethION, currently still only available in early access, is the “big brother” on the MinION. It’s a desktop-sized machine offering the promise of outputs that rival the largest, highest throughput machines of Illumina.
- Ultra-long reads
- Low capital costs
- Portability (MinION)
- Single molecule
The Oxford Nanopore system is primarily comprised of a nanopore (the current version of which, the “R9”, is based on the E. coli pore CsgG) embedded in an artificial membrane (version designated as “M#”) and a motor protein (version designated as “E#”) which helps translocate DNA molecules from one side of the membrane to the other through the nanopore. A voltage is applied across the membrane causing an electric current to run through the nanopore. As DNA passes through the nanopore it modifies the current, with different bases having slightly different effects. These differences are used to reconstruct the DNA sequence and, in some cases, base modifications as well. It works with single molecules, so it doesn’t suffer from amplification bias that is present in many other sequencers, although its error rate, currently at 5-10%, is substantially higher than that of market leader Illumina. The main error mode appears to be systematic homopolymer errors, although there is the promise that software can overcome this issue. It produces very long reads, with several reports of individual reads of several hundred kilobases long, making it ideal for de novo assembly of genomes and addressing regions of the genome with large repeats. One of the more interesting capabilities is the ability to directly sequence RNA molecules, eliminating the biases inherent in reverse transcription when converting RNA to DNA.
There has been a tremendous amount of interest in Oxford Nanopore’s sequencers and much discussion on how disruptive it will be to the overall sequencing market. It is difficult, however, to make direct comparisons with other sequencers. First, due to the nature of the technology and the way in which it is being rolled out by Oxford Nanopore, there have been rapid changes to the platform, with new pore versions coming out every few months. Also, the concept of a “run” doesn’t really apply to the platform the way it does for other sequencers. There is no set run time (although there appears to be a practical limit of about 48 hours) and no fixed read length, leading to completely tunable system outputs. There’s even the possibility of programming the system to sequence only desired stretches of DNA or avoiding unwanted stretches of DNA.
The low capital and running costs of the MinION have encouraged a large number of researchers to at least try out the platform, with over 3000 placed so far. Despite the broad base of users, coaxing the most out of the system still takes effort, skill and practice. Therefore, the platform is still finding its footing, with particular success in certain niches such as directly sequencing infectious disease samples in the field.
The MinION, Oxford Nanopore’s only fully released platform, is undergoing rapid improvements, leading to larger outputs, longer reads, and higher quality data. The PromethION is currently in early access and promises to offer sequencing outputs and costs that rival the largest, highest throughput machines of Illumina. The SmidgION is a future platform that will take portability to the next level, as it will plug in directly to a smartphone. They are also working on two automated sample prep platforms. The VolTRAX will help feed the PromethION while the Zumbador is being designed for field sample prep. Oxford Nanopore has also hinted at a number of other ambitious, far-reaching projects, including solid-state nanopores, protein sequencing, and even DNA synthesis.
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