An improved PCR-based method for site directed mutagenesis using megaprimers
Research output: Contribution to journal › Journal article › Research › peer-review
An improved protocol for site-directed mutagenesis based on the two-step polymerase chain reaction (PCR) megaprimer method is described. Compared to previously published protocols, the protocol described in this article ensures consistently a success rate of at least 85% with essentially no introduction of unwanted secondary mutations. The essential features of this protocol include an optimization of the template-primer amounts and ratio that allows the use of a reduced number of PCR cycles and the use of proof-reading thermostable DNA polymerases.
|Journal||Molecular and Cellular Probes|
|Number of pages||3|
|Publication status||Published - 1998|
Keywords: DNA Primers; DNA-Directed DNA Polymerase; Humans; Hydroxymethylbilane Synthase; Mutagenesis, Site-Directed; Point Mutation; Polymerase Chain Reaction