NANOPORE-BASED NUCLEIC ACID ANALYSIS WITH MIXED FRET DETECTION

摘要

Various methods, systems and devices for optical detection and analysis of polymers, such as polynucleotides, using nanopores, e.g., for determining sequences of nucleic acids, are provided herein. In certain variations, methods and systems for determining a nucleotide sequence of a polynucleotide, which include measuring mixed FRET signals as a polynucleotide translocates through a nanopore and determining a nucleotide sequence of the polynucleotide from the mixed FRET signals, are provided.

主权项

1. A method of determining a nucleotide sequence of a polynucleotide, the method comprising the steps of:
translocating a polynucleotide through a nanopore so that nucleotides of the polynucleotide pass in sequence by a first member of a FRET pair positioned adjacent to the nanopore, a plurality of the nucleotides being within a FRET distance of the first member of the FRET pair as the nucleotides exit the nanopore and at least a portion of the nucleotides being labeled with a second member of the FRET pair; exposing the FRET pairs adjacent to the nanopore to a light beam so that FRET occurs between the first and a plurality of second members of the FRET pair within the FRET distance to generate a mixed FRET signal; measuring mixed FRET signals as the polynucleotide translocates through the nanopore; and determining a nucleotide sequence of the polynucleotide from the mixed FRET signals.