US 2014/0350235 A1 · IN SITU OLIGONUCLEOTIDE SYNTHESIS ON A PARAMAGNETIC SUPPORT

Abstract

A novel method for attaching oligonucleotides to a paramagnetic solid support is disclosed. Conventional methods of attachment require that oligonucleotides be pre-synthesized with specific end modifications, which is laborious and expensive. Instead, we attached oligonucleotides to paramagnetic beads by direct synthesis of the oligonucleotides on the surface of the beads. An external magnet was used to hold the paramagnetic beads in place during solid-phase synthesis. A magnetic force was applied directly to the beads to prevent their loss, in particular, during reagent purge-to-waste steps that involved high-pressure drain or vacuum. This method can be adapted for use in any laboratory working with conventional synthesis automation, and can be employed, for example, with single columns and multi-well titer plates.

Claims48

Dependent claims

A method for synthesis of an oligonucleotide, the method comprising:

a) providing a paramagnetic solid support comprising a substrate coating the surface of the solid support, wherein the substrate comprises one or more functional groups capable of covalent attachment to a linker;
b) providing a magnet that attracts the paramagnetic solid support, whereby the paramagnetic solid support is immobilized on the surface of the magnet or a surface in contact with the magnetic field of the magnet;
c) covalently attaching phosphoramidite linkers to functional groups on the substrate to produce a derivatized support;
d) reacting the derivatized support with a nucleoside phosphoramidite corresponding to the first nucleotide of the desired oligonucleotide sequence such that the nucleoside phosphoramidite attaches covalently to a phosphoramidite linker;
and
e) adding nucleoside phosphoramidites stepwise to the growing nucleotide chain until an oligonucleotide having the desired sequence is produced.

3 more elements

Dependencies are inferred from the claim text.

In situ oligonucleotide synthesis on a paramagnetic support

Life of this patent

In situ oligonucleotide synthesis on a paramagnetic support

Life of this patent

Abstract

Claims48