Biopolymer Sequencing By Hybridization of probes to form ternary complexes and variable range alignment
Methods for sequencing a biopolymer by forming local ternary complexes along the length of the double-stranded biopolymer target molecule using one or more probes and obtaining information about the location of the probe(s) using a detector. These methods offer particular advantage when implemented with nanopore (including micropore) detection systems.
1. A method for determining a whole or partial sequence of a target biopolymer, the method comprising the steps of:
providing a double-stranded biopolymer target molecule;
contacting said target molecule or fragment thereof with a first probe having specificity for one or more first recognition sites of said target molecule, thereby forming one or more first local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof, said one or more first recognition sites each having a first known sequence, wherein the first probe is a member selected from the group consisting of an oligonucleotide, a peptide, and a polyamide;
contacting said target molecule or fragment thereof with a second probe having specificity for one or more second recognition sites of said target molecule, thereby forming one or more second local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof, said one or more second recognition sites each having a second known sequence;
passing said double-stranded biopolymer target molecule or fragment thereof through a detector configured to detect a ternary complex through an increase or decrease of signal;
detecting an electrical signal indicative of one or more locations of at least a subset of said one or more first local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof to determine a first probe map;
detecting an electrical signal indicative of one or more locations of at least a subset of said one or more second local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof to determine a second probe map; and
determining said whole or partial sequence of said target biopolymer using at least said first probe map and said second probe map.
2. A method for determining a whole or partial sequence of a target biopolymer, the method comprising the steps of:
providing a single-stranded molecule;
providing a double-stranded biopolymer target molecule by adding a second strand to the single-stranded molecule;
contacting said target molecule or fragment thereof with a plurality of n probes each having specificity for one or more recognition sites of said target molecule, thereby forming local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof, each of said recognition sites having a known sequence, wherein each of the n probes is individually a member selected from the group consisting of an oligonucleotide, a peptide, and a polyamide;
detecting at least a subset of said local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof to determine one or more probe maps; and
determining said whole or partial sequence of said target biopolymer using at least said one or more probe maps.
3. A method for determining a whole or partial sequence of a target biopolymer, the method comprising the steps of:
providing a double-stranded biopolymer target molecule;
contacting said target molecule or fragment thereof with a plurality of n probes each having specificity for one or more recognition sites of said target molecule, thereby forming local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof, each of said recognition sites having a known sequence, wherein each of the n probes is individually a member selected from the group consisting of an oligonucleotide, a peptide, and a polyamide;
passing said double-stranded biopolymer target molecule or fragment thereof through a detector configured to detect a ternary complex through an increase or decrease of signal;
detecting at least a subset of said local ternary complexes along said double-stranded biopolymer target molecule or fragment thereof to determine one or more probe maps; and
determining said whole or partial sequence of said target biopolymer using at least said one or more probe maps.