Nucleic acid analysis device, method for producing same, and nucleic acid analyzer

Abstract

Disclosed is a technique for binding microparticles to patterned bonding pads of a metal (e.g., gold) formed on a support. The microparticles each carry a nucleic acid synthetase or DNA probe immobilized thereon for capturing a nucleic acid sample fragment. The technique involves forming, on a support surface, a film having a thickness equivalent to that of the bonding pads; controlling the size of microparticles with respect to the size of bonding pads; and thereby immobilizing microparticles each bearing a single nucleic acid sample fragment to the bonding pads in a one-to-one manner in a grid form. This allows high-density regular alignment and immobilization of many types of nucleic acid fragment samples on a support and enables high-throughput analysis of nucleic acid samples. Typically, immobilization of microparticles at 1-micrometer intervals easily provides a high density of 10 6 nucleic acid fragments per square millimeter.

1 . A nucleic acid analysis device, comprising:
 a support;   a plurality of bonding pads arranged on a surface of the support;   a thin-film layer covering the surface of the support in a region other than the bonding pads;   microparticles, where a single microparticle is bound to each of the bonding pad; and   a probe molecule or molecules of a single type immobilized on each of the microparticle,   wherein the microparticles are bound to the bonding pads through chemical bonds, and wherein the thin-film layer is capable of suppressing non-specific adsorption of the microparticles on the surface of the support.   
     
     
         2 . The nucleic acid analysis device of  claim 1 , wherein a single probe molecule is immobilized on a single microparticle. 
     
     
         3 . The nucleic acid analysis device of  claim 1 , wherein the probe molecule or molecules comprise a nucleic acid or a nucleic acid synthetase. 
     
     
         4 . The nucleic acid analysis device of  claim 1 , wherein the microparticles is a material selected from the group consisting of semiconductors, metals, inorganic polymers, and organic polymers. 
     
     
         5 . The nucleic acid analysis device of  claim 1 ,
 wherein the bonding pads comprise a material selected from the group consisting of gold, titanium, nickel, and aluminum.   
     
     
         6 . A method for producing a nucleic acid analysis device which comprises the steps of:
 forming a thin metal film on a surface of a support;   selectively etching the thin metal film to form a plurality of bonding pads;   introducing a linear molecule into each of the bonding pad, the linear molecule capable of being adsorbed on the bonding pad;   binding an each of the microparticles to the linear molecule through a chemical bond; and   immobilizing a probe molecule or molecules to each of the microparticle through a chemical bond.   
     
     
         7 . The method for producing a nucleic acid analysis device of  claim 6 , wherein the bonding pads are regularly arranged on the support. 
     
     
         8 . The method for producing a nucleic acid analysis device of  claim 6 , wherein two or more probe molecules are immobilized on a single microparticle. 
     
     
         9 . The method for producing a nucleic acid analysis device of  claim 6 , wherein the bonding pads have diameters each twice or less the diameters of the microparticles. 
     
     
         10 . A method for producing a nucleic acid analysis device which comprises the steps of:
 forming a thin metal film on a surface of a support;   introducing a linear molecule film onto the thin metal film, the linear molecule film capable of being adsorbed on the thin metal film;   forming a plurality of dummy bonding pads, selectively etching the the linear molecule film and thin metal film, removing the dummy bonding pads to expose the linear molecule film, and thereby forming a plurality of bonding pads each having the thin metal film and the linear molecule film;   binding each of the microparticles to the exposed linear molecule film through a chemical bond; and   immobilizing a probe molecule or molecules to each of the microparticles through a chemical bond.   
     
     
         11 . The method for producing a nucleic acid analysis device of  claim 10 , wherein the bonding pads are regularly arranged on the support. 
     
     
         12 . The method for producing a nucleic acid analysis device of  claim 10 , wherein two or more probe molecules are immobilized on a single microparticle. 
     
     
         13 . The method for producing a nucleic acid analysis device of  claim 10 , wherein the bonding pads have diameters each twice or less the diameters of the microparticles. 
     
     
         14 . A nucleic acid analyzer which comprises:
 a nucleic acid analysis device including a support and microparticles regularly immobilized on a surface of the support, each of the microparticles having a probe molecule or molecules capable of capturing a nucleic acid to be analyzed;   a supplier for supplying a nucleic acid sample and a nucleotide having a fluorescent dye to the nucleic acid analysis device;   an irradiator for irradiating the nucleic acid analysis device with light; and   an emission detector for detecting fluorescence emitted from the fluorescent dye incorporated into a nucleic acid chain through nucleic acid elongation induced by the coexistence of a nucleotide, a nucleic acid synthetase, and a nucleic acid sample;   wherein the nucleic acid analyzer obtains information on nucleotide sequence of the nucleic acid sample, and   wherein the nucleic acid analysis device further comprises a plurality of bonding pads arranged on the surface of the support at positions where the microparticles are immobilized, the microparticles are bound to the bonding pads through chemical bonds, and the nucleic acid analyzer further comprises a linear molecule film covering the support in a region other than the bonding pads.   
     
     
         15 . The nucleic acid analyzer of  claim 14 , wherein a single probe molecule is immobilized on a single microparticle. 
     
     
         16 . The nucleic acid analyzer of  claim 14 , wherein the probe molecule or molecules comprise a nucleic acid or a nucleic acid synthetase. 
     
     
         17 . The nucleic acid analyzer of  claim 14 , wherein the microparticles comprise a material selected from the group consisting of semiconductors, metals, inorganic polymers, and organic polymers. 
     
     
         18 . The nucleic acid analyzer of  claim 14 , the bonding pads comprise a material selected from the group consisting of gold, titanium, nickel, and aluminum.