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

Abstract:

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.

Nucleic Acids Research.1997:25(11):2227

Lehrstuhl Biologie der Mikroorganismen, Ruhr-Universität, Universitätsstrasse 150, D

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