Microfabricated integrated dna analysis system

Abstract

Methods and apparatus for genome analysis are provided. A microfabricated structure including a microfluidic distribution channel is configured to distribute microreactor elements having copies of a sequencing template into a plurality of microfabricated thermal cycling chambers. A microreactor element may include a microcarrier element carrying the multiple copies of the sequencing template. The microcarrier element may comprise a microsphere. An autovalve at an exit port of a thermal cycling chamber, an optical scanner, or a timing arrangement may be used to ensure that only one microsphere will flow into one thermal cycling chamber wherein thermal cycling extension fragments are produced. The extension products are captured, purified, and concentrated in an integrated oligonucleotide gel capture chamber. A microfabricated component separation apparatus is used to analyze the purified extension fragments. The microfabricated structure may be used in a process for performing sequencing and other genetic analysis of DNA or RNA.

1 . A microfabricated structure comprising:
 a distribution channel to distribute microreactor elements carrying multiple copies of a clonal sequencing template into a plurality of thermal cycling chambers such that only one microreactor element will pass into one thermal cycling chamber wherein thermal cycling extension fragments are produced from a microreactor element;   purification chambers connected to the thermal cycling chambers to capture and concentrate the extension fragments; and   component separation channels connected to the purification chambers to analyze the extension fragments.   
     
     
         2 . The microfabricated structure of  claim 1  wherein the microreactor element includes a microcarrier element that carries the multiple copies of the clonal sequencing template. 
     
     
         3 . The microfabricated structure of  claim 1  wherein the microreactor element is a bolus or a microemulsion droplet. 
     
     
         4 . The microfabricated structure of  claim 3  wherein the microreactor element includes a microsphere carrying the multiple copies of the clonal sequencing template. 
     
     
         5 . The microfabricated structure of  claim 1  wherein the sequencing template is a DNA or RNA sequencing template. 
     
     
         6 . A system for performing sequencing comprising:
 means for shearing DNA or RNA into fragments;   means for ligating the fragments to form a mixture of desired circular and contaminating linear products;   means for selectively removing the contaminating linear products;   means for generating microreactor elements carrying multiple clonal copies of a single sequencing template;   means for selecting which microreactor elements have a sequencing template;   microfluidic distribution means for distributing a selected microreactor element with a sequencing template into a thermal cycling chamber;   means for ensuring that only one microreactor element will flow into one thermal cycling chamber;   extension means, including the thermal cycling chambers, for producing thermal cycling extension fragments from the microreactor elements carrying multiple copies of the sequencing template;   purification chamber means for capturing, purifying and concentrating the extension fragments; and   component separation means for analyzing the extension fragments.   
     
     
         7 . The system of  claim 6  wherein the microreactor element includes a microcarrier element that carries the multiple copies of the clonal sequencing template. 
     
     
         8 . The system of  claim 6  wherein the microreactor element is a bolus or a microemulsion droplet. 
     
     
         9 . The system of  claim 8  wherein the microreactor element includes a microsphere carrying the multiple copies of the clonal sequencing template. 
     
     
         10 . A microfabricated structure comprising:
 a distribution channel to distribute microspheres carrying multiple copies of a clonal sequencing template into a plurality of thermal cycling chambers;   an autovalve at an exit port of a thermal cycling chamber to ensure that only one microsphere will flow into one thermal cycling chamber wherein thermal cycling extension fragments are produced from a microsphere;   purification chambers connected to the thermal cycling chambers to capture and concentrate the extension fragments; and   component separation channels connected to the purification chambers to analyze the extension fragments.   
     
     
         11 . The microfabricated structure of  claim 10  wherein the diameter of a microsphere is between about 1 and 100 microns. 
     
     
         12 . The microfabricated structure of  claim 11  wherein the diameter of a microsphere is about 10 microns. 
     
     
         13 . The microfabricated structure of  claim 10  wherein each thermal cycling chamber is in fluid communication with a separation channel. 
     
     
         14 . The microfabricated structure of  claim 10  wherein the sequencing template is a DNA or RNA sequencing template. 
     
     
         15 - 44 . (canceled)