Method for the uncoupled, direct, exponential amplification and sequencing of DNA molecules with the addition of a second thermostable DNA polymerase and its application

US6828094B2 · kind B2 · utility

Assignee

• Roche Diagnostics GMBH · DE

Inventors

• Christian Kilger · Hersbruck, DE
• Svante Paabo · Hersbruck, DE
• Michael Motz · Heidelberg, DE

Key dates

• Filing date: Jun 24, 1999
• Grant date: Dec 7, 2004
• Expiry date: Jun 24, 2019

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12Q1/6869
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

Method for sequencing a nucleic acid molecule in a thermocycling reaction which initially comprises a nucleic acid molecule, a first primer, a second primer, a reaction buffer, a first thermostable DNA polymerase, (optionally) a thermostable pyrophosphatase, deoxynucleotides or derivatives thereof and a dideoxynucleotide or a derivative thereof and which is characterized in that the thermocycling reaction additionally contains a second thermostable DNA polymerase which, in comparison to the said first thermostable DNA polymerase, has a reduced ability to incorporate dideoxynucleotides as well as the use of the said method.