Nucleotide analogues

• US 10,301,346 B2
• Filed: 11/04/2016
• Issued: 05/28/2019
• Est. Priority Date: 11/06/2015
• Status: Active Grant

• Current Assignee: IsoPlexis Corporation (Bruker Corporation)
• Original Assignee: QIAGEN Sciences, LLC (Qiagen NV)
• Inventors: Marma, Mong Sano, Olejnik, Jerzy, Korboukh, Ilia
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US15/343,267
• Publication Number:
  US 20170130051A1
• Time in Patent Office: 935 Days
• Field of Search: 536 231, 536 266, 536 41
• CPC Class Codes: C07C 2603/18 Fluorenes; Hydrogenated fluorenes
• CPC Class Codes: C07C 323/12 the carbon skeleton being acyclic and saturated
• CPC Class Codes: C07C 323/60 with the carbon atom of at least one of the carboxyl groups bound to nitrogen atoms
• CPC Class Codes: C07D 207/46 with hetero atoms directly attached to the ring nitrogen atom
• CPC Class Codes: C07F 7/1804 Compounds having Si-O-C linkages Si-O-acyl linkages C07F7/1896
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 19/14 Pyrrolo-pyrimidine radicals
• CPC Class Codes: C09B 1/00 Dyes with anthracene nucleus not condensed with any other ring
• CPC Class Codes: C09B 11/24 Phthaleins containing amino groups ; Phthalanes; Fluoranes; Phthalides; Rhodamine dyes; Phthaleins having heterocyclic aryl rings; Lactone or lactame forms of triarylmethane dyes
• CPC Class Codes: C09B 23/083 five >CH- groups
• CPC Class Codes: C09B 23/166 containing two or more nitrogen atoms hydrazon dyes -CH=N-N- C09B26/02
• CPC Class Codes: C09B 5/2436 only nitrogen-containing hetero rings
• CPC Class Codes: C12Q 1/6811 Selection methods for production or design of target specific oligonucleotides or binding molecules
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: Y02P 20/55 Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Abstract

The present invention provides methods, compositions, mixtures and kits utilizing deoxynucleoside triphosphates comprising a 3′-O position capped by a group comprising methylenedisulfide as a cleavable protecting group and a detectable label reversibly connected to the nucleobase of said deoxynucleoside. Such compounds provide new possibilities for future sequencing technologies, including but not limited to Sequencing by Synthesis.

Claims

1. A deoxynucleoside triphosphate having the structure:

2. The deoxynucleoside triphosphate according to claim 1, wherein said nucleobase is non-natural.

3. The deoxynucleoside triphosphate according to claim 2, wherein the non-natural nucleobase is selected from the group consisting of 7-deaza guanine, 7-deaza adenine, 2-amino,7-deaza adenine, and 2-amino adenine.

4. The deoxynucleoside triphosphate according to claim 1, wherein said cleavable protecting group is of the formula —

CH₂—

SS—

R, wherein R is selected from the group consisting of alkyl and substituted alkyl groups.

5. The deoxynucleoside triphosphate according to claim 1, wherein said detectable label is a fluorescent label.

9. The deoxynucleoside triphosphate according to claim 1, wherein L₁ is selected from the group consisting of—

CONH(CH₂)_x—

, —

CO—

O(CH₂)_x—

, —

CONH—

(OCH₂CH₂O)_x—

, —

CO—

O(CH₂CH₂O)_x—

, and —

CO(CH₂)_x—

, wherein x is 0-10.

10. The deoxynucleoside triphosphate according to claim 1, wherein L₂ is selected from the group consisting of

6. A labeled deoxynucleoside triphosphate according to the following structure:

7. The labeled deoxynucleoside triphosphate according to claim 6, wherein L₂ is not —

S—

.

8. A deoxynucleoside triphosphate having the structure: