Characterization of individual polymer molecules based on monomer-interface interactions
View Patent ↗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. A polymer analysis system comprising:
(a) a structure comprising an ion-permeable passage connecting a first pool and a second pool, wherein a polymer to be analyzed is placed in the first pool;
(b) a molecular motor disposed adjacent to the ion-permeable passage, wherein the molecular motor is a processive nucleic acid translocation molecule, wherein said processive nucleic acid translocation molecule is not a bacteriophage receptor capable of transporting bacteriophage DNA; and
(c) a detection system operative to detect a property of a target polymer moving relative to the ion-permeable passage.
2. The polymer analysis system of claim 1 , wherein a protein comprises said ion-permeable passage.
3. The polymer analysis system of claim 2 , wherein the molecular motor is fused to the protein.
4. The polymer analysis system of claim 1 , wherein the structure comprises a lipid bilayer.
5. The polymer analysis system of claim 1 , wherein the translocation molecule comprises a DNA polymerase, a RNA polymerase, an exonuclease, or a ribosome.
6. The polymer analysis system of claim 5 , wherein the DNA polymerase is an E. coli DNA polymerase I or a T7 DNA polymerase.
7. The polymer analysis system of claim 5 , wherein the RNA polymerase is T7 RNA polymerase.
8. The polymer analysis system of claim 5 , wherein the exonuclease is exonuclease Lambda.
9. The polymer analysis system of claim 1 , wherein the molecular motor is disposed on the side of the structure cis to the first pool.
10. The polymer analysis system of claim 1 , wherein the molecular motor is disposed on the side of the structure trans to the first pool.
11. The polymer analysis system of claim 1 , wherein the ion permeable passage has a diameter of about 1.0 to 4.0 nanometers.
12. The polymer analysis system of claim 1 , wherein the ion-permeable passage has a diameter of about 0.5 to 2 nanometers.
13. The polymer analysis system of claim 1 , wherein the detection system is operative to detect an electrical property of the target polymer.