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US 2010/0143900 A1Published application
Asynchronous sequencing of biological polymers
AssigneePacific Biosciences of Californ, Inc.
InventorsPaul S. Peluso, John S. Eid, David Rank
Life of this patent
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Abstract
The present invention provides methods, reagents and apparatus for conducting single molecule sequencing in an asynchronous manner. The subject methods and compositions are particularly useful for high throughput and multiplexed sequencing.
Claims20
A method for sequencing a target nucleic acid molecule, the method comprising:
- (a) providing a first nucleic acid molecule and a second nucleic acid molecule, the first nucleic acid molecule and the second nucleic acid molecule having the same or essentially the same sequences as the target nucleic acid molecule, wherein the first nucleic acid molecule is complexed with a first primer and a first polymerase and the second nucleic acid molecule is complexed with a second primer and a second polymerase;
- (b) subjecting the first nucleic acid molecule to a first polymerization reaction to yield a first strand complementary to the first nucleic acid molecule;
- (c) subjecting the second nucleic acid molecule to a second polymerization reaction to yield a second strand complementary to the second nucleic acid molecule;
- (d) establishing asynchronous polymerization reactions between the first nucleic acid molecule and the second nucleic acid molecule;
- (e) obtaining time sequences of incorporation of nucleotides or nucleotide analogs into the first strand and the second strand during the asynchronous polymerization reactions;
- and
- (f) sequencing the target nucleic acid molecule by compiling the time sequences of incorporation.
4 more elements
A method for sequencing a target nucleic acid molecule, the method comprising:
- (a) providing two or more nucleic acid molecules having the same or essentially the same sequences as the target nucleic acid molecule, wherein each of the two or more nucleic acid molecules is complexed with a primer and a polymerase, wherein individual polymerases are complexed at different locations of the target nucleic acid molecule such that at least two nascent strands of different but overlapping sequences are achieved;
- (b) subjecting the two or more nucleic acid molecules to polymerization reactions to yield the at least two nascent strands;
- (c) obtaining time sequences of incorporation of nucleotides or nucleotide analogs into individual nascent strands during the polymerization reactions;
- and
- (d) compiling the time sequences of incorporation, thereby sequencing the target nucleic acid molecule.
2 more elements
A method for sequencing a target nucleic acid molecule, the method comprising:
- (a) providing two or more nucleic acid molecules having the same or essentially the same sequences as the target nucleic acid molecule, wherein each of the two or more nucleic acid molecules is complexed with a primer and a polymerase;
- (b) subjecting the two or more nucleic acid molecules to polymerization reactions to yield at least two nascent strands of different but overlapping sequences;
- (c) exposing the two or more nucleic acid molecules to light for an exposure time that is less than a length of time required to form a full-length strand complementary to the target nucleic acid molecule using the polymerase, the full-length strand and the target nucleic acid molecule having the same or essentially the same number of nucleotides;
- (d) obtaining time sequences of incorporation of nucleotides or nucleotide analogs into individual nascent strands during the polymerization reactions;
- and
- (e) compiling the time sequences of incorporation, thereby sequencing the target nucleic acid molecule.
3 more elements
An apparatus for sequencing a target nucleic acid molecule, the apparatus comprising:
- an array of optical confinements for sequencing nucleic acid molecules, the optical confinements comprising target nucleic acid molecules;
- and
- a computer system configured to compile time sequences of incorporation of nucleotides or nucleotide analogs during asynchronous polymerization of two or more nucleic acid strands having different but overlapping sequences complementary to each of the target nucleic acid molecules.
Dependencies are inferred from the claim text.