Methods of nucleic acid analysis using terminator nucleotides
First Claim
1. A method for detecting a nucleotide incorporation, comprising:
- a) providing a surface having a plurality of reaction sites, wherein each reaction site is attached or operatively linked to at least one sensor, and one or more of the reaction sites contain (i) a polymerase, (ii) a nucleic acid template and (iii) an extendible end;
b) contacting the reaction sites with a first solution containing one or more types of terminator nucleotides, wherein the one or more types of terminator nucleotides comprise (1) a terminator moiety and (2) a tag moiety comprising a first binding partner that is capable of binding to a second binding partner that comprises a signal-generation moiety;
c) incorporating at least one type of a terminator nucleotide at an extendible end and generating a non-extendible end having a terminator moiety, thereby generating a nucleotide incorporation product within one or more of the reaction sites;
d) contacting the first binding partner of a tag moiety of the terminator nucleotide in the nucleotide incorporation product with a second binding partner comprising a signal-generation moiety thereby binding the first binding partner with the second binding partner; and
e) detecting a non-optical signal generated by the signal-generation moiety of the second binding partner at one or more of the reaction sites using the sensors thereby detecting a nucleotide incorporation at the one or more reaction sites, wherein the signal is detected after incorporation of the nucleotide.
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Accused Products
Abstract
In some embodiments, the disclosure relates generally to methods, as well as related, systems, compositions, kits and apparatuses, for nucleic acid analysis that involve the use of modified nucleotides, including terminator nucleotides and/or tagged nucleotides, in a template-dependent nucleotide incorporation reaction. In some embodiments, the nucleic acid analysis can be conducted at a single reaction site, or at a plurality of reaction sites in an array of reaction sites. Optionally, the array contains a plurality of reaction sites having about 1-100 million, or about 100-250 million, or about 200-500 million, or about 500-900 million, or more reaction sites. Optionally, each reaction site is in contact with, operatively coupled, or capacitively coupled to one or more sensors that are ion-sensitive FETs (isFETs) or chemically-sensitive FETs (chemFETs) sensors. Optionally, the reaction sites are in fluid communication with each other.
34 Citations
20 Claims
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1. A method for detecting a nucleotide incorporation, comprising:
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a) providing a surface having a plurality of reaction sites, wherein each reaction site is attached or operatively linked to at least one sensor, and one or more of the reaction sites contain (i) a polymerase, (ii) a nucleic acid template and (iii) an extendible end; b) contacting the reaction sites with a first solution containing one or more types of terminator nucleotides, wherein the one or more types of terminator nucleotides comprise (1) a terminator moiety and (2) a tag moiety comprising a first binding partner that is capable of binding to a second binding partner that comprises a signal-generation moiety; c) incorporating at least one type of a terminator nucleotide at an extendible end and generating a non-extendible end having a terminator moiety, thereby generating a nucleotide incorporation product within one or more of the reaction sites; d) contacting the first binding partner of a tag moiety of the terminator nucleotide in the nucleotide incorporation product with a second binding partner comprising a signal-generation moiety thereby binding the first binding partner with the second binding partner; and e) detecting a non-optical signal generated by the signal-generation moiety of the second binding partner at one or more of the reaction sites using the sensors thereby detecting a nucleotide incorporation at the one or more reaction sites, wherein the signal is detected after incorporation of the nucleotide. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20)
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Specification