Nature Methods | Methagora

Next-next DNA sequencing

While the technology feature, “DNA sequencing: generation next-next”, was at press, Pacific Biosciences of Menlo Park, California stunned the community with their announcement of a single molecule sequencing technology they claim will provide a complete human genome in 15 minutes by the year 2013. Although Pacific Biosciences was founded in 2004, the company had been very ‘hush hush’ about their technology development. But that veil of secrecy was lifted during the Advances in Genome Biology and Technology meeting held February 6th to 9th at Marco Island, Florida where Stephen Turner, chief technology officer, presented the first preliminary data on the system.

According the company’s website, their single molecule sequencing method, called SMRT for ‘single molecule, real time’, is based on a zero mode waveguide which helps create very small detection volumes. The reaction is performed in a well that is 70 nanometers wide and holds only 20 zeptoliters (10E-21 liters) of detection volume occupied by a single molecule of polymerase. The key to the technology is illuminating around the polymerase, but not above, as it incorporates each fluorescently tagged nucleotide into an elongating strand of DNA. The remaining nucleotides are above the polymerase, in the dark, thereby greatly reducing background fluorescence which has been an issue for other developers. As each nucleotide is incorporated by the polymerase, its fluorescence is detected by a CCD camera and the fluorescent tag is cleaved, leaving the elongating DNA in a native form.

With a described sequencing rate of 10 bases per second, in real-time, performed across thousands of these wells on the chip, the Pacific Biosciences instrument would provide faster sequencing than any other machine available today. While they hope to have an instrument ready for 2010, they are targeting 2013 for the arrival of a model that can deliver the stunning 15-minute genome. The company is still working to reach that target by increasing the number of zero mode waveguides on a chip to one million and modifying the polymerase to increase incorporation to 50 nucleotides per second. With these enhancements they are projecting a mind boggling 100 gigabases/hour.

So, what is your opinion of all the effort going into ‘next-next generation’ sequencing technology and the stir that it has created? And if a ‘15 minute genome’ becomes reality — what are the first things that should be done using such a technology?

Nathan Blow, Technology Editor


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    Roald Forsberg said:

    Over at CLC bio we are thinking a lot about these issues. Next generation sequencing techniques are upon us and are creating great bioinformatics challenges. However, in a few years next-next generation technologies will arrive and we need to get ready for this already. The two main challenges, as we see it, are: firstly the data handling and analysis challenge. How do we keep up with an instrument that can potentially create 100 Gb/hour ? Our answer to this is algorithm optimization and the integration of accelerated algorithms through the use of SIMD and FPGA technology. Secondly, the filtering, analysis and ultimate presentation of all the sequencing information to the end users. As sequencing costs drop and novel application areas are explored, the end users of a DNA sequencing analysis will no longer be only specialized geneticists and researchers, but equally likely be technicians, GP’s of lay people. The answer to this is less trivial and requires a lot of interaction and dialog with end users such as clinicians and molecular biologists.

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    Nathan Blow said:

    I wonder if sequencing is going to find itself in a similar situation as in mass spec, where the instrumentation is now far ahead of the software. But I guess we will find out very quickly — Helicos shipped their first single molecule sequencer on Wednesday, which they say has the imaging capacity to interrogate approximately 1.3 billion bases per hour. This is going to be the first look at single molecule sequencing as a commercial product. And while not 100GB/hr yet, Helicos says that there is tremendous headroom on the system to increase output and, as this post shows, the other single molecule sequencing companies are moving fast. It is probably only a matter of time.

    The issue of getting this information into the hands of the GP’s for their patients is a very complicated one. Although this might be solved with good software and education of GP’s and patients (along the lines of what the PGP is proposing), I was very surprised to hear from a number of people that they would not want their genomes sequenced even if it could be done quickly and cheaply.