Saturation Mutagenesis by Mutagenic Oligonucleotide-Directed PCR Amplification (Mod-PCR)

Saturation mutagenesis is one approach for determining the contributions of individual base pairs or amino acids to the structure and function of defined DNA sequence elements or proteins, respectively (1 ). If a simple phenotypic screen is available, saturation mutagenesis can be used to determine the relative importance of regions (larger than single basepairs or amino acids) by dividing a large element into smaller regions and independently saturating each small region with random point mutations. The number and degree of phenotypic changes exhibited by each regionally distinct population of mutants indicate the importance of each region to the activity of the whole element (2 ).