Roche Diagnostics and 454 launch next generation sequencer

By Dr Matt Wilkinson

- Last updated on GMT

Related tags Dna sequencing Dna

The first next generation sequencer from Roche and 454 is already
proving popular only weeks after its release.

Roche Diagnostics has announced the worldwide launch its Genome Sequencer FLX System, developed in collaboration with CuraGen subsidiary, 454 Life Sciences.

The first Genome Sequencer FLX systems were delivered to evaluation sites in November 2006, including major genome centers.

Now, only two weeks after its launch, the companies are already benefiting from the first sales.

The system is capable of expressed sequence tag (EST) sequencing, the identification of small RNAs and transcription binding sites as well as whole genome sequencing and metagenomics.

The instrument improves on the previous Genome Sequencer 20 system as it has been shown to give longer read lengths, a crucial feature for drug developers.

Talking to LabTechnologist.com, Dr Mary Shramke, 454's vice president of marketing, said: "One of the things that is really important for high throughput DNA sequencing is for people to see that researchers can look at the whole genome of bacteria and viruses."

"It's a really powerful way to understand how organisms evolve and adapt, to see how bacteria mutate to develop drug resistance, for example, and you have to do that on the DNA level," she continued.

The new technology can also complete a run of 100 mega bases (MB) in less than eight hours.

Commenting on the new release, Dr Shramke said: "There may be other platforms that can measure more bases in a run, but you need to ask how long do the runs take and how accurate are the results."

The technology is based on 'sequencing-by-synthesis' technology, which begins by breaking up the DNA sample into fragments that are between 300 and 500 bases in length.

"You get better confidence with the longer strands, they are crucial for putting the sequence together when overlap becomes quite a significant issue; longer reads are so important in getting a very high consensus accuracy," continued Dr Shramke.

These fragments are then ligated by two short adaptors, which provide primers for both amplification and sequencing of the fragment as well as a biotin tag that immobilises it onto a streptavidin-coated bead.

Very dilute solutions containing the DNA fragments are used, such that each 28um bead binds only a single DNA fragment.

The beads are then emulsified with polymerase chain reaction (PCR) reagents in water-in-oil microreactors so that clonal amplification can occur.

This yields beads with millions of copies of the DNA fragment attached.

These beads are then deposited in 454's PicoTiterPlate device by centrifugation before enzyme beads and incubation mix are added.

The PicoTiterPlate contains over 1.6m wells that are approximately 44um wide, so only fit one bead per well.

Each well has an optical fibre attached to its base, which form an array leading to the astronomy grade CCD camera.

The fluidics sub-system allows nucleotides to be pumped in, in a fixed order.

During the nucleotide flow each of the beads is sequenced in parallel, with the polymerase extending the sequencing strand only if the nucleotide is complimentary to the template strand.

The addition of one (or more) nucleotides results in a reaction that generates a light signal, which is recorded by the instrument's camera.

Dr Shramke said: "If you sequence more than one nucleotide at once you get a stronger signal and we see a very linear response between the signal strength and the number of nucleotides added."

"We will be looking at working towards the $100,000 genome sequence landmark in 2007."

Any potential customers using the old technology need not worry about a complicated upgrade process either.

"GS 20 systems already in use can be upgraded easily to the Genome Sequencer FLX on-site," said Manfred Baier, head of Roche applied science.

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