Most cystic fibrosis (CF) patients can blame their malady on asingle gene mutation, but more than 300 other mutated genes alsoclaim a piece of the CF action. To identify them all for purposes ofdiagnosis and possible therapy is a formidable proposition at best.But a marriage of biomolecular and electronic technologiespromises to speed up the sequencing and synthesis ofoligonucleotides many-fold, for many applications, including CFgene analysis.That promise is being made by Affymetrix, of Santa Clara, Calif.,the diagnostics subsidiary of Palo Alto, Calif.-based Affymax NV.Its latest expression is in today's issue of the Proceedings of theNational Academy of Sciences (PNAS). "Light-generatedoligonucleotide arrays for rapid DNA sequence analysis" is the titleof the paper by senior author Stephen P. Fodor, Affymetrix'sscientific director.What's involved is using the technology by which Silicon Valleymakes semiconductor chips, to create miniaturized, densely packednucleic-acid arrays. "It's a little like reducing the size of a 96-wellmicrotiter plate to the dimensions of a pin-head, or smaller," Fodortold BioWorld Today.His "sequence analysis by hybridization" method uses a set of shortoligo probes of defined sequence to seek out complementarysequences on a longer target DNA strand.To begin, his team coats a silica support with oligonucleotidegroups tethered to glass by their 3'. The 5' ends are protected byphoto-sensitive chemical groups that come off when exposed tofrequencies of light in the near-violet 365-nanometer range. Thesurface-bound molecule can then couple to the next incomingnucleoside.A microscopic matrix of transparent and opaque squares serves as amask, with openings as small as 20 square microns. These lightpulses illuminate a pre-set pattern, Fodor explained, "just asphotolithography etches semiconductor chip surfaces to defineelectronic circuits." This degree of miniaturization is limited onlyby the diffraction wavelength of light.It's still necessary to amplify the sequence under analysis, butinstead of the conventional gel-based resequencing of a kilobase ofDNA at a time, and electrophoresis of labeled DNA fragments, theDNA-chip technology goes straight to hybridization.For example, Fodor said, "instead of one probe at a time, the chiparray could sequence an entire exon of the CF gene at once, makinggenetic screening of patients and potential parental carriers morepractical."Compared to current methods, he added, "the technique takes one totwo hours instead of all day."Since submitting their progress report to PNAS last January, Fodorsaid, "we've made a lot of progress. For instance, in our paper wereported producing a half-inch square array of 256 different octa-nucleotides in 16 chemical reaction cycles, completed in four hours.Since then, "we've gone from 256 components to well over50,000."
-- David N. Leff Science Editor
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