Comparative Analysis of Three Site-directed Mutagenesis Methods

Objective: Analysis of improved three site-directed mutagenesis methods in a novel gene recombinants construction. Methods: Overlapping extension PCR, MutanBest kit and Quikchange site-directed mutagenesis kit were used to construct mutants con-taining a single different residue from wild type gene. By Stratagagene primers design on-line, PCR was simplified. Comparing with PrimeSTAR polymerase and Ultra-Super competent cell kit substitution, Quikchange kit protocol was more economical. Results: Objec-tive recombinants were successfully gained by all three methods, so recombinant vectors could be utilized in further test. Conclusions: With two key elements substitution, Quikchang site-directed mutagenesis protocol could be improved to an effective, convenient, simple and economic approach.