Chip-based sequencing nucleic acids

Abstract

A system for fast DNA sequencing by amplification of genetic material within microreactors, denaturing, demulsifying, and then sequencing the material, while retaining it in a PCR/sequencing zone by a magnetic field. One embodiment includes sequencing nucleic acids on a microchip that includes a microchannel flow channel in the microchip. The nucleic acids are isolated and hybridized to magnetic nanoparticles or to magnetic polystyrene-coated beads. Microreactor droplets are formed in the microchannel flow channel. The microreactor droplets containing the nucleic acids and the magnetic nanoparticles are retained in a magnetic trap in the microchannel flow channel and sequenced.

The invention claimed is: 
     
         1 . A method of sequencing nucleic acids on a microchip, comprising the steps of:
 providing a microchannel flow channel in the microchip;   isolating the nucleic acids;   hybridizing the nucleic acids to magnetic nanoparticles or to magnetic polystyrene-coated beads;   forming microreactors in said microchannel flow channel, said microreactors containing the nucleic acids and said magnetic nanoparticles or magnetic polystyrene-coated beads;   positioning said microreactors containing the nucleic acids and said magnetic nanoparticles or magnetic polystyrene-coated beads in a magnetic trap in said microchannel flow channel, and   sequencing the nucleic acids.   
     
     
         2 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of hybridizing the nucleic acids to magnetic nanoparticles or to magnetic polystyrene-coated beads comprises hybridizing the nucleic acids to magnetic nanoparticles. 
     
     
         3 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of hybridizing the nucleic acids to magnetic nanoparticles or to magnetic polystyrene-coated beads comprises hybridizing the nucleic acids to magnetic-cored optically discrete nanoparticles. 
     
     
         4 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of hybridizing the nucleic acids to magnetic nanoparticles or to magnetic polystyrene-coated beads comprises hybridizing the nucleic acids to magnetic polystyrene-coated beads. 
     
     
         5 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of sequencing the nucleic acids comprises pyrosequencing the nucleic acids. 
     
     
         6 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of sequencing the nucleic acids comprises four-color sequencing the nucleic acids by synthesis. 
     
     
         7 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein the nucleic acids being analyzed include virions, capsids, or individual cells containing the nucleic acids and wherein said step of isolating the nucleic acids comprises lysing the virions, capsids, or individual cells containing the nucleic acids by acoustic wave lysing, or by heating, or by optical heating, or by chemical lysing, or by laser-induced lysing, followed by hybridizing to the magnetic beads. 
     
     
         8 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of positioning said microreactors containing the nucleic acids and said magnetic nanoparticles or magnetic polystyrene-coated beads in a magnetic trap in said microchannel flow channel comprises moving the nucleic acids in a hydrophobic carrier fluid to said magnetic trap. 
     
     
         9 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of positioning said microreactors containing the nucleic acids and said magnetic nanoparticles or magnetic polystyrene-coated beads in a magnetic trap in said microchannel flow channel comprises forcing the nucleic acid into a water-oil emulsification hydrophobic carrier fluid and moving the nucleic acid in said water-oil emulsification hydrophobic carrier fluid to said magnetic trap. 
     
     
         10 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of positioning said microreactors containing the nucleic acids and said magnetic nanoparticles or magnetic polystyrene-coated beads in a magnetic trap in said microchannel flow channel comprises forcing the nucleic acid into liquid slugs in a carrier fluid and moving the nucleic acid in said liquid slugs to said magnetic trap. 
     
     
         11 . The method of sequencing nucleic acids on a microchip of  claim 1  including the step of flowing reagent over the nucleic acids in said magnetic trap. 
     
     
         12 . The method of sequencing nucleic acids on a microchip of  claim 1  wherein said step of providing a microchannel flow channel in the microchip comprises providing a microchannel flow channel cross section aspect ratio, width and depth, that prevents said magnetic nanoparticles or magnetic polystyrene-coated beads from vertical stacking. 
     
     
         13 . The method of sequencing nucleic acids on a microchip of  claim 1  including the step of placing a cover over said microchip during said steps of isolating the nucleic acids;
 hybridizing the nucleic acids to magnetic nanoparticles or to magnetic polystyrene-coated beads; 
 positioning said microreactors containing the nucleic acids and said magnetic nanoparticles or magnetic polystyrene-coated beads in a magnetic trap in said microchannel flow channel, and 
 sequencing the nucleic acids. 
 
     
     
         14 . A method of fast DNA sequencing by amplification of genetic material within microreactors, comprising the steps of:
 providing a microchannel flow channel,   hybridizing the genetic material to magnetic nanoparticles or to magnetic polystyrene-coated beads,   forming the microreactors in said microchannel flow channel, and   retaining the genetic material in a PCR/Sequencing Zone by a magnetic field.   
     
     
         15 . The method of fast DNA sequencing by amplification of genetic material within microreactors of  claim 14  wherein the method encounters reaction products or excess reagents, including the step of holding the genetic material in said PCR/Sequencing Zone by said magnetic field and washing away the reaction products or the excess reagents or removing the reaction products or the excess reagents by chemical degradation. 
     
     
         16 . The method of fast DNA sequencing by amplification of genetic material within microreactors of  claim 14  including the step of turning off the magnetic field permitting the genetic material to be flushed from said PCR/Sequencing Zone to waste or to archival storage.