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

1997; Oxford University Press; Volume: 25; Issue: 11 Linguagem: Inglês

10.1093/nar/25.11.2227

1362-4962

Andreas Urban,

RNA Interference and Gene Delivery

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.