Controlling oligonucleotide surface density in light-directed DNA array fabrication.

Over the past two decades high-density DNA arrays have developed into a central technology for nucleic acid analyses. Important application areas include whole-genome gene expression studies, high throughput analyses of single nucleotide polymorphisms, and, most recently, the determination of binding site specificities for transcription factors and other critical elements involved in gene regulation. A key parameter in the performance of DNA arrays is the density of the surface-bound oligonucleotides, which strongly affects both thermodynamic and kinetic aspects of DNA hybridization. In this report, we describe an approach for the control of oligonucleotide density in photolithographically fabricated DNA arrays, based upon a controlled UV light deprotection procedure. Modulation of the UV exposure permits a desired degree of deprotection of surface synthesis sites; a subsequent capping reaction to inactivate the exposed sites leaves only a desired fraction of active sites remaining for synthesis, corresponding to a lower oligonucleotide density. It is shown that the procedure is reasonably general, in that it is readily transferable to alternative substrate materials with similar results.