Random Mutagenesis by Whole-Plasmid PCR Amplification

Mutagenesis is a popular tool used in the analysis of protein structure and function. Polymerase chain reaction (PCR)-based mutagenesis can be used to introduce mutations with the use of the appropriate primer. Although the majority of attention has been given to site-directed mutagenesis, random mutagenesis is actually an older approach and has considerable potential because of its limited bias, if an appropriate screening method is available. This approach has successfully been used to obtain “gain-of-function” mutants (1 ). The ability to target mutants to individual proteins and parts of proteins with modern molecular tools has considerable applicability.