Fiberoptic Bead Array

A new ultra HT method now used during the generation of the forthcoming haplotype map is an assay taking advantage of fiberoptic bead arrays developed by Illumina®. This technology takes advantage of fiberoptic array bundles and specifically prepared 3-mm sensor beads with specific DNA probe (address or ZIP code) sequences immobilized on the surface of each bead (Fig. 5B).

Up to 1520 different ZIP oligo sequences are pooled in one tube. Fibers are then introduced in bead pools, where beads self-assemble into pits at the fiber end. The assembly of beads is then checked by excitation of the fibers that transmit photons by internal reflection, where the photon either leaves the fiber end or is emitting fluorescence if hitting an attached bead. The bead that is self-assembled to a particular fiber is then tested by a defined series of hybridization and washing steps, making it possible to guess the present oligo ZIP code.

DNA template is prepared in the meantime by an allele-specific extension and ligation assay that builds a sequence consisting of a universal primer, 5'-primer, extended genomic region, 3'-primer, ZIP code oligo, and another universal primer. This initial step is then followed by a PCR using the two universal primers, leading to a molecule that can be probed with the fiber bundle for the presence of the ZIP code oligo.

The arrays are fabricated into microplate-compatible array matrices containing 16, 96, or 384 separate arrays. Illumina's 96-bundle array matrix can process up to 150,000 assays in parallel in a single experiment, making more than 1 million assays possible per day. Due to the probing of a predefined oligo, accuracy seems to be better than in assays that have to detect any variable genomic sequence.

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