Multiple orthogonal labelling of oligonucleotides

US9315537B2 · kind B2 · utility

Assignees

• Deutsches Krebsforschungszentrum · DE
• RUPRECHT - KARLS - UNIVERSITÄT HEIDELBERG · DE

Inventors

• Juliane Becker · Mannheim, DE
• Andres Jäschke · Heidelberg, DE
• Ayan Samanta · Mannheim, DE
• Manfred Wiessler · Bischofswerda, DE

Key dates

• Filing date: Aug 31, 2012
• Grant date: Apr 19, 2016
• Expiry date: Sep 24, 2032

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C07H21/00
• WIPO field: Organic fine chemistry
• WIPO sector: Chemistry

Abstract

In summary, the present invention concerns a method for multiple orthogonal labelling of oligonucleotides, preferably RNA or DNA, by simultaneously performing the inverse Diels-Alder reaction (DAinv) and the copper-catalyzed click reaction (CuAAC), wherein the method is employed in a single step by just adding the different reaction components together and incubating the aqueous reaction mixture preferably for one hour at room temperature. In detail, the reaction components are one or more N3-modified labels, a copper compound, a stabilizing ligand, a reducing agent and one or more electron-deficient label-modified dienes that are added together with an at least double-modified oligonucleotide having one more nucleotides containing one or more N3-reactive groups and one or more electron-rich dienophiles, wherein a terminal alkyne moiety is preferably used as N3-reactive group(s) and a frans-cyclooctene moiety or norbornene is preferably used as electron-rich dienophile(s), more preferably frans-cyclooctene. Therefore, the present invention provides a one-pot method for post-synthetic multiple orthogonal labeling of oligonucleotides, which allows the site-specific introduction of mo…