Direct cloning of PCR amplified nucleic acids

US5487993A · kind A · utility

Assignees

• Invitrogen Corporation · US
• Chimer · US

Inventors

• Corinna Herrnstadt · San Diego, US
• Joseph Fernandez · Carlsbad, US
• Lloyd M. Smith · Atlanta, US
• David A. Mead · Madison, US

Key dates

• Filing date: Sep 9, 1993
• Grant date: Jan 30, 1996
• Expiry date: Sep 9, 2013

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12Q1/6853
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

Methods are described for producing recombinant DNA molecules from suitable host vectors and nucleic acids subjected to 3'-terminal transferase activity. In one embodiment, the method takes advantage of the single 3'-deoxy-adenosine monophosphate (dAMP) residues attached to the 3' termini of PCR generated nucleic acids. Vectors are prepared with recognition sequences that afford single 3'-terminal deoxy-thymidine monophosphate (dTMP) residues upon reaction with a suitable restriction enzyme. Thus, PCR generated copies of genes can be directly cloned into the vectors without need for preparing primers having suitable restriction sites therein. The invention also contemplates associated plasmid vectors and kits for implementing the methods.