Characterization of individual polymer molecules based on monomer-interface interactions

Abstract

The invention relates to a method for detecting a double-stranded region in a nucleic acid by (1) providing two separate, adjacent pools of a medium and a interface between the two pools, the interface having a channel so dimensioned as to allow sequential monomer-by-monomer passage of a single-stranded nucleic acid, but not of a double-stranded nucleic acid, from one pool to the other pool; (2) placing a nucleic acid polymer in one of the two pools; and (3) taking measurements as each of the nucleotide monomers of the single-stranded nucleic acid polymer passes through the channel so as to differentiate between nucleotide monomers that are hybridized to another nucleotide monomer before entering the channel and nucleotide monomers that are not hybridized to another nucleotide monomer before entering the channel.

1 - 11 . (canceled)  
   
   
       12 . A method for determining the duration of interaction of a polymer with a nanopore, the method comprising the steps of: 
 (a) providing a structure comprising a nanopore connecting a first pool and a second pool, wherein a polymer to be analyzed is placed in the first pool;    (b) causing the polymer to traverse the ion permeable passage; and    (c) measuring the duration of step (b).    
   
   
       13 . The method of  claim 12 , wherein the structure further comprises a molecular motor.  
   
   
       14 . The method of  claim 12 , wherein a protein comprises the nanopore.  
   
   
       15 . The method of  claim 12 , wherein the structure comprises a lipid bilayer.  
   
   
       16 . The method of  claim 13 , wherein the molecular motor comprises a DNA polymerase, a RNA polymerase, an exonuclease, or a ribosome.  
   
   
       17 . The method of  claim 16 , wherein the DNA polymerase is an  E. coli  DNA polymerase I or a T7 DNA polymerase.  
   
   
       18 . The method of  claim 16 , wherein the RNA polymerase is T7 RNA polymerase.  
   
   
       19 . The method of  claim 16 , wherein the exonuclease is exonuclease Lambda.  
   
   
       20 . The method of  claim 12 , wherein the nanopore has a diameter of about 1.0 to 4.0 nanometers.  
   
   
       21 . The method of  claim 12 , wherein the nanopore has a diameter of about 0.5 to 2 nanometers.  
   
   
       22 . The method of  claim 12 , wherein the polymer comprises a polynucleotide.  
   
   
       23 . The method of  claim 12 , wherein the structure further comprises a DNA binding protein.  
   
   
       24 . The method of  claim 12 , wherein the measuring detects an electrical property of the structure while the polymer traverses the nanopore.  
   
   
       25 . The method of  claim 12 , wherein a protein does not comprise the nanopore.