Methods and apparatus that increase sequencing-by-binding efficiency
First Claim
1. A method of nucleic acid detection, comprising steps of:
- (a) contacting a primed template nucleic acid with a polymerase and a first mixture of nucleotides under conditions for forming a ternary complex at a nucleotide position in the template, wherein the first mixture comprisesa nucleotide cognate of a first base type comprising a label that produces a first signal;
a nucleotide cognate of a second base type comprising a label that produces the first signal;
a nucleotide cognate of a third base type comprising a label that produces a second signal; and
a nucleotide cognate of a fourth base type comprising a label that produces the second signal;
(b) contacting the primed template nucleic acid with a polymerase and a second mixture of nucleotides under conditions for forming a ternary complex at the nucleotide position in the template, wherein the second mixture comprisesa nucleotide cognate of the first base type comprising a label that produces the first signal;
a nucleotide cognate of a second base type comprising a label that produces the second signal;
a nucleotide cognate of a third base type comprising a label that produces the second signal; and
a nucleotide cognate of the fourth base type comprising a label that produces the first signal; and
(c) examining products of steps (a) and (b) for signals produced by a ternary complex that comprises the primed template nucleic acid, a polymerase and a next correct nucleotide.
3 Assignments
0 Petitions
Accused Products
Abstract
A method of nucleic acid detection including (a) contacting a primed template with a polymerase, nucleotide cognate of a first base type and nucleotide cognate of a second base type under ternary complex stabilizing conditions; (b) contacting the primed template nucleic acid with a polymerase, nucleotide cognate of the first base type and nucleotide cognate of a third base type under ternary complex stabilizing conditions; (c) examining products of (a) and (b) for signals produced by a ternary complex that includes the primed template nucleic acid, polymerase and next correct nucleotide, wherein signals for the product of (a) are ambiguous for the first and second base type, and signals for the product of (b) are ambiguous for the first and third base type; (d) disambiguating signals acquired in (c) to identify the next correct nucleotide.
-
Citations
23 Claims
-
1. A method of nucleic acid detection, comprising steps of:
-
(a) contacting a primed template nucleic acid with a polymerase and a first mixture of nucleotides under conditions for forming a ternary complex at a nucleotide position in the template, wherein the first mixture comprises a nucleotide cognate of a first base type comprising a label that produces a first signal; a nucleotide cognate of a second base type comprising a label that produces the first signal; a nucleotide cognate of a third base type comprising a label that produces a second signal; and a nucleotide cognate of a fourth base type comprising a label that produces the second signal; (b) contacting the primed template nucleic acid with a polymerase and a second mixture of nucleotides under conditions for forming a ternary complex at the nucleotide position in the template, wherein the second mixture comprises a nucleotide cognate of the first base type comprising a label that produces the first signal; a nucleotide cognate of a second base type comprising a label that produces the second signal; a nucleotide cognate of a third base type comprising a label that produces the second signal; and a nucleotide cognate of the fourth base type comprising a label that produces the first signal; and (c) examining products of steps (a) and (b) for signals produced by a ternary complex that comprises the primed template nucleic acid, a polymerase and a next correct nucleotide. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 13, 14, 15, 16, 17)
-
-
10. A method of nucleic acid detection, comprising:
-
(a) sequentially contacting a primed template nucleic acid with first and second mixtures under conditions for forming a ternary complex, wherein each of the mixtures comprises a polymerase and nucleotide cognates for at least two of four different base types in the primed template nucleic acid, wherein the first mixture comprises a nucleotide cognate of a first base type comprising a label that produces a first signal; a nucleotide cognate of a second base type comprising a label that produces the first signal; a nucleotide cognate of a third base type comprising a label that produces a second signal; and a nucleotide cognate of a fourth base type comprising a label that produces the second signal; and
the second mixture comprisesa nucleotide cognate of the first base type comprising a label that produces the first signal; a nucleotide cognate of a second base type comprising a label that produces the second signal; a nucleotide cognate of a third base type comprising a label that produces the second signal; and a nucleotide cognate of the fourth base type comprising a label that produces the first signal; (b) examining the first and second mixtures, or products thereof, separately to detect ternary complexes; and (c) identifying the next correct nucleotide for the primed template nucleic acid molecule, wherein the next correct nucleotide is identified as a cognate of one of the four different base types if ternary complex is detected in the two mixtures. - View Dependent Claims (11, 12, 18, 19, 20, 21, 22, 23)
-
Specification