Ultrafast sequencing of biological polymers using a labeled nanopore

Abstract

Methods and systems for sequencing a biological molecule or polymer, e.g., a nucleic acid, are provided. One or more donor labels, which are attached to a pore or nanopore, may be illuminated or otherwise excited. A polymer having a monomer labeled with one or more acceptor labels, may be translocated through the pore. Either before, after or while the labeled monomer of the polymer passes through, exits or enters the pore, energy may be transferred from the excited donor label to the acceptor label of the monomer. As a result of the energy transfer, the acceptor label emits energy, and the emitted energy is detected in order to identify the labeled monomer of the translocated polymer and to thereby sequence the polymer.

1 - 16 . (canceled) 
     
     
         17 . A method for sequencing nucleic acid polymers comprising:
 translocating nucleic acid polymers each in single file through a plurality of nanopores, wherein the nucleic acid polymers each comprise nucleotide monomers labeled with acceptor labels and the nanopores each have attached one or more donor labels each capable of fluorescence resonanant energy transfer (FRET) after illumination with radiation and wherein acceptor labels come in close proximity with donor labels as nucleotide monomers exit the nanopores so that a FRET energy exchange interaction can occur between donor labels and acceptor labels;   exciting donor labels by illumination so that energy from excited donor labels is transferred by FRET to the acceptor labels of nucleotide monomers exiting the nanopores, wherein such energy transfer causes the acceptor labels to emit radiation at a lower energy than the radiation that was used to excite the donor labels;   detecting the radiation emitted by the acceptor labels as a result of the energy transfer, wherein the radiation emitted by the acceptor labels identify the nucleotide monomer to which it is attached; and   determining nucleotide sequences of the nucleic acid polymers based on the detection of the emitted radiation from the acceptor labels and identifications of the nucleotide monomers.   
     
     
         18 . The method of  claim 17  wherein said nucleic acid polymers are single stranded nucleic acids. 
     
     
         19 . The method of  claim 18  wherein said step of translocating said single stranded nucleic acids through said nanopores comprises applying an electric field through the nanopores. 
     
     
         20 . The method of  claim 18  wherein said one or more donor labels are selected from the group consisting of quantum dots, metal nanoparticles, nanodiamonds and fluorophores. 
     
     
         21 . The method of  claim 20  wherein said one or more donor labels comprise quantum dots. 
     
     
         22 . The method of  claim 18  wherein said acceptor labels are fluorophores. 
     
     
         23 . The method of  claim 18  wherein said plurality of nanopores comprises a nanopore membrane assembly and wherein said nanopores are protein nanopores. 
     
     
         24 . The method of  claim 23  wherein said step of determining nucleotide sequences utilizes both optical and electrical readouts from said nanopore membrane assembly. 
     
     
         25 . The method of  claim 18  wherein in said step of translocating said acceptor labels come within 10 nm of donor labels as said nucleotides exit said nanopores. 
     
     
         26 . The method of  claim 18  wherein in said step of translocating said acceptor labels come within a Förster radius of a donor label as said nucleotides exit said nanopores. 
     
     
         27 . The method of  claim 18  wherein a plurality of said donor labels exhibiting different spectral emission maxima are attached to the nanopores and a plurality of acceptor labels having distinct absorption/emission spectra each associated with a specific nucleotide monomer are attached to said single stranded nucleic acid, and wherein each donor label and corresponding acceptor label undergo a FRET (Förster Resonance Energy Transfer) interaction such that each acceptor label emits a distinct spectra by which its associated nucleotide monomer is identified. 
     
     
         28 . The method of  claim 27  wherein said plurality of nanopores comprises a nanopore membrane assembly and wherein said nanopores are protein nanopores. 
     
     
         29 . The method of  claim 27  wherein in said step of translocating said acceptor labels come within 10 nm of donor labels as said nucleotides exit said nanopores. 
     
     
         30 . The method of  claim 27  wherein in said step of translocating said acceptor labels come within a Forster radius of donor labels as said nucleotides exit said nanopores. 
     
     
         31 . The method of  claim 18  wherein said step of detecting said radiation emitted by said acceptor labels comprises optical detection of distinct wavelengths emitted by different acceptor labels each associated with a specific nucleotide monomer. 
     
     
         32 . The method of  claim 31  wherein said plurality of nanopores comprises a nanopore membrane assembly and wherein said nanopores are protein nanopores. 
     
     
         33 . The method of  claim 31  wherein in said step of translocating said acceptor labels come within 10 nm of donor labels as said nucleotides exit said nanopores. 
     
     
         34 . The method of  claim 31  wherein in said step of translocating said acceptor labels come within a Förster radius of donor labels as said nucleotides exit said nanopores. 
     
     
         35 . The method of  claim 18  wherein at least one of four nucleotide monomers are labeled with said acceptor labels and time dependent signals arising from sequences of emissions from nucleotide monomers are converted into nucleic acid sequences corresponding to positions of the at least one of four labeled nucleotides in the single stranded nucleic acids translocating through said plurality of nanopores. 
     
     
         36 . The method of  claim 35  wherein the method is repeated for different at least one of the four nucleotide monomers in separate samples and time dependent signals therefrom are deduced and then aligned to assemble an entire nucleic acid sequence. 
     
     
         37 . The method of  claim 35  wherein said plurality of nanopores comprises a nanopore membrane assembly and wherein said nanopores are protein nanopores. 
     
     
         38 . The method of  claim 35  wherein in said step of translocating said acceptor labels come within 10 nm of donor labels as said nucleotides exit said nanopores. 
     
     
         39 . The method of  claim 35  wherein in said step of translocating said acceptor labels come within a Förster radius of donor labels as said nucleotides exit said nanopores.