A new method for attaching oligodeoxyribonucleotides to glass involving monoalkoxylated and dialkoxylated silanes and bromoacetamide/phosphorothioate linking chemistry has been developed. Three novel bromoacetamide silanes were synthesized for derivatization of glass microscope slides by traditional dipping methods. A thin film method that conserves silane and provides a consistent protocol for test experiments was also used. Oligonucleotides bearing 5′-phosphorothioates were synthesized by literature methods. Immobilization conditions were initially established by treatment of bromoacetamidosilyl slides with fluoresceinated oligonucleotides, which were imaged by confocal fluorescence microscopy. Spotting can be accomplished in water at oligonucleotide concentrations down to 0.1 mM. Oligonucleotides immobilized using this method can serve as primers for templated, polymerase-based extension reactions with a fluoresceinated dideoxynucleotide terminator. When such primers are formatted into small arrays, specific extension is observed only in the presence of complementary template, with the amount of immobilized primer reflected in the fluorescence signal.

A high fidelity, surface-based method of nucleic acid analysis has been developed based on DNA polymerase extension of primer-template complexes on DNA microchips. The ability of the method to discriminate against mismatches and provide an almost "digital" signal recommended it for molecular computation. A DNA computer with the capability of solving nondeterministic polynomial time (NP)-complete problems (those whose time-complexity function rises exponentially with the problem size) in polynomial time using this Arrayed Primer EXtension (APEX) method was experimentally demonstrated. An algorithm involving extension of surface-bound primer-template complexes, representing solutions and clauses of a Boolean formula, is described for the solution of two-, three-, and four-variable satisfiability (SAT) problems, including a 3SAT, exploiting the theoretical concepts of Lipton. A discussion of the principles of nondeterministic computing with APEX is also provided.

No abstract