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Nucleic Acids Research, Vol 25, Issue 11 2227-2228, Copyright © 1997 by Oxford University Press

ARTICLES

A rapid and efficient method for site-directed mutagenesis using one- step overlap extension PCR

A Urban, S Neukirchen and KE Jaeger
Lehrstuhl Biologie der Mikroorganismen, Ruhr-Universitat, Universitatsstrasse 150, D-44780 Bochum, Germany.

A rapid method is described to efficiently perform site-directed mutagenesis based on overlap extension polymerase chain reaction (OE- PCR). Two template DNA molecules in different orientations relative to only one universal primer were amplified in parallel. By choosing a high dilution of mutagenic primers it was possible to run an overlap extension PCR in only one reaction without purification of intermediate products. This method which we have named one-step overlap extension PCR (OOE-PCR) can in principle be applied to every DNA fragment which can be cloned into a multiple cloning site of any common cloning vector.
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