Processing polynucleotide-containing samples

Abstract

Methods and systems for processing polynucleotides (e.g., DNA) are disclosed. A processing region includes one or more surfaces (e.g., particle surfaces) modified with ligands that regain polynucleotides under a first set of conditions (e.g., temperature and pH) and release the polynucleotides under a second set of conditions (e.g., higher temperature and/or more basic pH). The processing region can be used to, for example, concentrate polynucleotides of a sample and/or separate inhibitors of amplification reactions from the polynucleotides. Microfluidic devices with a processing region are disclosed.

1 - 23 . (canceled) 
     
     
         24 . A diagnostic apparatus configured to detect polynucleotides in a sample solution, the apparatus comprising:
 a first processing region comprised of a plurality of magnetic binding particles, the binding particles having polycationic polyamide ligands bound to the surfaces thereof, wherein the binding particles are configured to preferentially bind polynucleotides in the sample solution;   a second processing region comprising one or more one or more lyophilized particles containing one or more amplification reagents; and   an amplification region comprising:
 an inlet; 
 a microfluidic chamber; 
 an outlet; and 
 an inlet valve and an outlet valve, the inlet and outlet valves configured to isolate the sample solution in the amplification chamber during amplification. 
   
     
     
         25 . The apparatus of  claim 24 , further comprising a detection region in which the presence of amplified polynucleotides may be detected. 
     
     
         26 . The apparatus of  claim 25 , wherein the detection region comprises:
 an inlet;   a microfluidic chamber;   an outlet; and   an inlet valve and an outlet valve, the inlet and outlet valves configured to isolate the sample solution in the amplification chamber during amplification.   
     
     
         27 . The apparatus of  claim 24 , further comprising a lysing region comprising a heating element configured to lyse cells in the sample solution. 
     
     
         28 . The apparatus of  claim 24 , further comprising one or more amplification heating elements in thermal communication with the microfluidic chamber of the amplification chamber. 
     
     
         29 . The apparatus of  claim 24 , further comprising one or more separation heating elements in thermal communication with the first processing region to assist in the release of the polynucleotides from the binding particles. 
     
     
         30 . The apparatus of  claim 24 , wherein the polycationic polyamide ligands comprise at least one of poly-DL-ornithine, poly-L-lysine, and poly-D-lysine. 
     
     
         31 . The apparatus of  claim 24 , further comprising a lysing reagent reservoir. 
     
     
         32 . The apparatus of  claim 24 , further comprising a waste container. 
     
     
         33 . The apparatus of  claim 24 , wherein the inlet and outlet valve of the amplification region are in an initially open state prior to introduction of the sample solution into the microfluidic chamber of the amplification region. 
     
     
         34 . The apparatus of  claim 24 , further comprising a plurality of gates, wherein the gates are in an initially closed state. 
     
     
         35 . The apparatus of  claim 24 , further comprising a hydrophobic vent, wherein the vent comprises a layer of porous hydrophobic material. 
     
     
         36 . The apparatus of  claim 24 , wherein the binding particles have a collective volume of less than 5 microliters. 
     
     
         37 . The apparatus of  claim 24 , wherein the binding particles have an average diameter of between 4 microns and about 20 microns. 
     
     
         38 . The apparatus of  claim 24 , further comprising one or more actuators configured to supply fluidic pressure to the sample solution to move the sample solution through one or more microfluidic channels of the apparatus.