Site-Specific Mutagenesis

Definition

This is a process in which a single base in a piece of DNA is changed to a chemically modified version and the biological effects (e.g., mutation) are measured in living cells, with prokaryotes or eukaryotes.

Discussion

Site-specific mutagenesis is an approach to interrogating the mutagenic potential of a particular DNA lesion by placing it at a specific site in a fragment of DNA, inserting this vector into a cell, and then measuring the mutagenic events that occur (Fig. 1). (The process should not be confused with site-directed mutagenesis, in which individual amino acids of a protein are systematically replaced.) The method was first reported in 1984 in Prof. John Essigmann’s laboratory (Loechler et al 1984), who constructed a vector containing O ⁶-methylguanine and then demonstrated its mutagenicity in Escherichia coli. Following replication of the bacteria, the mutants can be screened using several methods, including a certain phenotype (if the mutation leads to a...