N.sup.4 -substituted cytidines and process for incorporation into an oligonucleotides

• US 5,405,950 A
• Filed: 10/09/1987
• Issued: 04/11/1995
• Est. Priority Date: 05/15/1985
• Status: Expired due to Term

• Current Assignee: Amoco Corporation (BP PLC)
• Original Assignee: Amoco Corporation (BP PLC)
• Inventors: Mock, Graham A., Lovern, Douglas H.
• Primary Examiner(s): Rollins, John W.
• Assistant Examiner(s): CRANE, LAWRENCE E
• Application Number:
  US07/108,843
• Time in Patent Office: 2,741 Days
• Field of Search: 536/27, 536/28, 536/29, 536/25.32, 536/25.33, 536/25.34, 536/27.14, 536/28.50, 435/6, 935/77, 935/78
• US Class Current: 536/25.32
• CPC Class Codes: C07H 19/06 Pyrimidine radicals
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids

Abstract

An oligonucleotide containing at least one cytidine analog labeled at C4 with a reporter group.

Claims

1. A modified nucleoside having the formula ##STR6## in which X is a linker containing between 1 and 18 carbon atoms inclusive;

Y is --S-- or --CO.O--;

Z is S-benzyl, S-p-methoxybenzyl, S-p-nitrobenzyl, S-4-picolyl, S-2-picolyl N-Oxide, S-9-anthrylmethyl, S-diphenylmethyl, S-di(p-methoxyphenyl)methyl, S-triphenylmethyl, S-2,4-dinitrophenyl, S-t-butyl, S-isobutoxymethyl hemithioacetal, S-2-tetrahydropyranyl hemithioacetal, S-acetamidomethyl, S-aminothioacetal, S-cyanomethyl, S-2-nitro-1-phenylethyl, S-2,2-bis(carboethoxy)ethyl, S-benzoyl, S-(N-ethylcarbamyl), and S-ethyl disulfide when Y is --S--; and

methyl, methoxymethyl, methylthiomethyl, tetrahydropyranyl, benzyloxymethyl, phenacyl, N-phthalimidomethyl, 2,2,2-trichloroethyl, 2-haloethyl, 2-(p-toluenesulfonyl)ethyl, t-butyl, cinnamyl, benzyl, triphenylmethyl, bis(o-nitrophenyl)methyl, 9-anthrylmethyl, 2-(9,10-dioxo)anthrylmethyl, piperonyl, trimethylsilyl, t-butyldimethylsilyl, S-t-butyl, 2-alkyl-1,3-oxa-3-olines, N,N-dimethylamide, N-7-nitroindoylamide, hydrazide, N-phenylhydrazide, and N,N-diisopropylhydrazide when Y is --CO.O--;

and

each R₁ and R₂ is --OH, --SH, or --H;

provided that at least one of R₁ and R₂ must be --OH or --SH.

2. The modified nucleoside of claim 1, wherein X is --(CH₂)_n with n being 3.

3. The modified nucleoside of claim 1, wherein Y is --S--.

4. The modified nucleoside of claim 1, wherein Y is --CO.O--.

5. A process for synthesizing an oligonucleotide which includes the repeated steps of joining a predetermined number of nucleosides via a phosphate bridge between their sugar moieties, wherein the improvement comprises using a modified nucleoside of claim 1 in at least one of the joining steps.

6. The process of claim 5 wherein the improvement further comprising removing Z from said modified nucleoside after all of said predetermined number of nucleosides have been joined together and subsequently attaching a detectable reporter group to Y to replace Z of said modified nucleoside.

7. The process of claim 5, wherein X is --(CH₂)_n with n being 3.

8. The process of claim 5, wherein Y is --S--.

9. The process of claim 5, wherein Y is --CO.O--.

10. The process of claim 8, wherein said modified nucleoside is used in the last joining step.

11. The process of claim 9, wherein said modified nucleoside is used in the last joining step.