PREPARATION OF OLIGORIBONUCLEOTIDES

• US 3,850,749 A
• Filed: 06/12/1972
• Issued: 11/26/1974
• Est. Priority Date: 06/16/1971
• Status: Expired due to Term

• Current Assignee: Gabriel Kaufmann, Uriel Z. Littauer
• Original Assignee: Gabriel Kaufmann, Uriel Z. Littauer
• Inventors: Kaufmann, Gabriel, Littauer, Uriel Z.
• Primary Examiner(s): Tanenholtz, Alvin E.
• Application Number:
  US05/262,143
• Time in Patent Office: 897 Days
• Field of Search: 195/28N
• US Class Current: 435/91.3
• 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
• CPC Class Codes: C12P 19/34 Polynucleotides, e.g. nucleic acids, oligoribonucleotides

Abstract

A process for the preparation of oligoribonucleotides of definend predetermined sequence containing the natural 3'"'"''"'"'-5'"'"''"'"'internucleotide linkage, which comprises reacting an oligonucleotide with a monofunctional substrate in the presence of polynucleotide phosphorylase, to form a product which is longer, by one monomeric unit than the initial oligonucleotide, isolating the product from the reaction mixture, removing the blocking agent and repeating the process a desired number of times, adding by each step a desired unit until the desired product of predetermined length and sequence is obtained.

Claims

1. A PROCESS FOR THE PREPARATION OF OLIGORIBONUCLEOTIDES OF DEFINED AND PREDETERMINED SEQUENCE CONTAINING THE NATURAL 3'"'"''"'"'-5'"'"''"'"'-INNTERNUCLEOTIDE LINKAGE, WHICH COMPRISES:

(A) REACTING DESIRED OLIGONUCLEOTIDE WITH A 2'"'"''"'"'(3'"'"''"'"')-O-ACYL NUCLEOSIDE DIPHOSPHATE IN THE PRESENCE OF POLYNUCLEOTIDE PHOSPHORYLASE, (B) ISOLATING THE PRODUCT FROM THE REACTION MIXTURE, AND (C) REMOVING THE 2'"'"''"'"'(3'"'"''"'"')-O-ACYL BLOCKING GROUP.

2. A process as claimed in claim 1, characterized in that the polynucleotide phosphorylase is used in watersoluble form.

3. A process as claimed in claim 1, characterized in that the polynucleotide phosphorylase is used in carrier-bound insoluble form.

4. A process as claimed in claim 1, wherein the 2'"'"''"'"'(3'"'"''"'"')-O-acylnucleoside diphosphate is a 2'"'"''"'"'(3'"'"''"'"')-O-acyl ester of ADP, GDP, UDP, CDP, IDP, XDP.

5. A process as claimed in claim 1, wherein the 2'"'"''"'"' 3'"'"''"'"'(-O-acyl blocking group is a propionyl, butyryl, valeryl, isobutyryl, isovaleryl or methoxybenzoyl group.

6. A process as claimed in claim 1, wherein the reaction, isolation and removal steps (a), (b) and (c) are repeated, in sequence, utilizing as the initial reactant the product isolated in the aforesaid step (b), to produce an oligoribonucleotide of any predetermined length and sequence.