The proliferation of large-scale DNA-sequencing projects in recent years has driven a search for alternative methods to reduce time and cost. With a 100-fold increase in throughput over current sequencing technology, the advance in sequencing opens up new uses for sequencing, including personalised medicine and diagnostics.
Analysts believe that this sequencing technique, which leverages the power of microfabrication, will get more powerful and cheaper each year as the technology continues to advance and miniaturise.
The researchers illustrated the technique by sequencing the genome of the Mycoplasma genitalium bacterium repeatedly in four hours, with up to and exceeding 99.99 per cent accuracy.
The technology is described in the paper: "Genome sequencing in microfabricated high-density picolitre reactors," in the July 31, 2005, online issue of Nature.'
Here they describe a scalable, highly parallel sequencing system with raw throughput significantly greater than that of state-of-the-art capillary electrophoresis instruments.
The apparatus uses a novel fibre-optic slide of individual wells and is able to sequence 25 million bases, at 99 per cent or better accuracy, in one four-hour run. An approximately 100-fold increase was achieved in throughput over current Sanger sequencing technology.
The researchers had developed an emulsion method for DNA amplification and an instrument for sequencing by synthesis using a pyrosequencing protocol optimised for solid support and picolitre-scale volumes.
The study additionally demonstrated the throughput and accuracy of this system by shotgun sequencing and de novo assembly of the Mycoplasma genitalium genome with 96 per cent coverage at 99.96 per cent accuracy in one run of the machine.
Francis Collins, Director of the National Human Genome Research Institute, pointed out the limits of similar technology, commenting on how 454's new technique would provide immediate solutions.
"It is clear that sequencing technology needs to continue to become smaller, faster and less expensive in order to fulfil the promise of personalised medicine."
Christopher McLeod, President and Chief Executive Officer of 454 Life Sciences said: "Because this technology is so affordable and easy to use, we are able to 'democratise' whole genome research, making it available to researchers outside of the major genome sequencing centres, which we believe will further speed the course to personalised medicine."
DNA sequencing has changed the nature of biomedical research and medicine. Reductions in the cost, complexity and time required to sequence large amounts of DNA, including improvements in the ability to sequence bacterial and eukaryotic genomes, will have significant scientific, economic and cultural impact.
Large scale sequencing projects, including whole-genome sequencing, have usually required the cloning of DNA fragments into bacterial vectors, amplification and purification of individual templates, followed by Sanger sequencing using fluorescent chain-terminating nucleotide analogues and either slab gel or capillary electrophoresis.
Current estimates put the cost of sequencing a human genome between $10 million and $25 million. Alternative sequencing methods have been described, however, no technology has displaced the use of bacterial vectors and Sanger sequencing as the main generators of sequence information.
Future increases in throughput, and a concomitant reduction in cost per base, may come from the continued miniaturisation of the fibre-optic reactors, allowing more sequence to be produced per unit area-a scaling characteristic similar to that which enabled the prediction of significant improvements in the integrated circuit at the start of its development cycle.