An improved PCR-based method for site directed mutagenesis using megaprimers

Research output: Contribution to journalJournal articleResearchpeer-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.
Original languageEnglish
JournalMolecular and Cellular Probes
Issue number6
Pages (from-to)345-8
Number of pages3
Publication statusPublished - 1998
Externally publishedYes

Bibliographical note

Keywords: DNA Primers; DNA-Directed DNA Polymerase; Humans; Hydroxymethylbilane Synthase; Mutagenesis, Site-Directed; Point Mutation; Polymerase Chain Reaction

ID: 11254093