Nucleotide analogs

Patent metadata

Field	Value
Publication number	US-9932623-B2
Application number	US-201414463412-A
Country	US
Kind code	B2
Filing date	Aug 19, 2014
Priority date	Aug 19, 2013
Publication date	Apr 3, 2018
Grant date	Apr 3, 2018

Abstract

Provided herein is technology relating to the manipulation and detection of nucleic acids, including but not limited to compositions, systems, methods, and kits related to nucleotides comprising 3′-O-propargyl deoxynucleotides wherein the 3′-O-propargyl deoxynucleotides are linked to a nucleic acid by a polymerase.

First claim

We claim:

1. A reaction mixture comprising one or more dNTPs, one or more 3′-O-propargyl nucleotide analogs, and a polymerase.

2. The reaction mixture of claim 1 further comprising a target nucleic acid.

3. The reaction mixture of claim 1 wherein the ratio of dNTPs to 3′-O-propargyl nucleotide analogs is from 1:500 to 500:1.

4. The reaction mixture of claim 1 wherein said polymerase is a polymerase from a Thermococcus species 9° N-7 and comprising one or more mutations selected from D141A, E143A, Y409V, and A485L.

5. The reaction mixture of claim 1 comprising a primer comprising an index or a barcode.

6. The reaction mixture of claim 1 comprising a target nucleic acid that is an amplicon.

7. A next generation sequencing (NGS) library comprising a plurality of nucleic acids, wherein each nucleic acid comprises a 3′-O-propargyl-nucleotide.

8. The NGS library of claim 7 wherein each nucleic acid of the plurality of nucleic acids comprises a subsequence of a target nucleic acid sequence.

9. The NGS library of claim 7 wherein each nucleic acid of the plurality of nucleic acids is conjugated to an oligonucleotide by a triazole linkage.

10. The NGS library of claim 7 comprising a plurality of 3′-O-propargyl-dN terminated nucleic acids.

11. The NGS library of claim 7 wherein each nucleic acid of the plurality of nucleic acids comprises an index or a barcode.

12. The NGS library of claim 7 wherein each nucleic acid of the plurality of nucleic acids comprises an adaptor oligonucleotide.

13. A NGS library produced by a method comprising synthesizing a plurality of nucleic acids using a mixture of dNTPs and one or more 3′-O-propargyl nucleotide analogs.

14. The NGS library of claim 13 wherein the molar ratio of dNTPs to 3′-O-propargyl nucleotide analogs is from 1:500 to 500:1.

15. The NGS library of claim 13 wherein the synthesis is performed by a Thermococcus species 9° N-7 and comprising one or more mutations selected from D141A, E143A, Y409V, and A485L.

16. The NGS library of claim 13 comprising a plurality of nucleic acids, wherein each nucleic acid of the plurality of nucleic acids comprises a subsequence of a target nucleic acid sequence.

17. The NGS library of claim 13 comprising a plurality of nucleic acids, wherein each nucleic acid of the plurality of nucleic acids is conjugated to an oligonucleotide by a triazole linkage.

18. The NGS library of claim 13 comprising a plurality of 3′-O-propargyl-dN terminated nucleic acids.

19. The NGS library of claim 13 comprising a plurality of nucleic acids, wherein each nucleic acid of the plurality of nucleic acids comprises an index or a barcode.

20. The NGS library of claim 13 comprising a plurality of nucleic acids, wherein each nucleic acid of the plurality of nucleic acids comprises an adaptor oligonucleotide.

Assignees

• Abbott Molecular Inc

Inventors

• Kim Dae Hyun

Classifications

• C12N15/1093

  General methods of preparing gene libraries, not provided for in other subgroups · CPC title

• C12Q1/6869

  Methods for sequencing · CPC title

• C07H19/10 Primary

  with the saccharide radical esterified by phosphoric or polyphosphoric acids · CPC title

• C07H19/20 Primary

  with the saccharide radical esterified by phosphoric or polyphosphoric acids · CPC title

• C07H21/04

  with deoxyribosyl as saccharide radical · CPC title