Solid phase methods for polynucleotide production

US7482119B2 · kind B2 · utility

Assignee

• Blue Heron Biotechnology, Inc. · US

Inventors

• Hsing-Yeh Parker · Woodinville, US
• John T. Mulligan · Seattle, US

Key dates

• Filing date: Oct 13, 2004
• Grant date: Jan 27, 2009
• Expiry date: Oct 13, 2024

Classification

• Technology area (CPC B): Performing Operations; Transporting
• CPC primary: B01J2219/00722
• WIPO field: Organic fine chemistry
• WIPO sector: Chemistry

Abstract

Polynucleotides having in excess of 1,000 nucleotides can be prepared using a solid phase synthesis technique. A feature of the technique is the use of a reusable solid support that contains covalently bound oligonucleotide. This covalently bound oligonucleotide is annealed to a bridge oligonucleotide, where the bridge is also annealed to a first oligonucleotide that forms a portion of the target polynucleotide. After the target polynucleotide is synthesized, it can be removed from the solid support under denaturing conditions, and the solid support re-used to prepare additional target polynucleotides. The yield of the target polynucleotide increases when shearing force is applied to the solid support that is linked to the growing oligonucleotide. This shearing force is thought to extend the growing end of the oligonucleotide away from contact with other oligonucleotide bound to the solid support and make that end more accessible to annealing with solution oligonucleotide. The synthesis is conveniently accomplished on a porous frit, where reagents and washing solutions are pumped through the frit.