ABSTRACT
The roles of specific nucleotides in a DNA or RNA molecule, or specific amino acids within a protein can be explored by altering the corresponding DNA sequence by site-directed mutagenesis. Also, random mutagenesis techniques are increasingly being used to explore biomolecule function. PCR provides an excellent tool for site-directed mutagenesis, including the deletion or insertion of sequences, the alteration of one or a few specific nucleotides, and the random mutation of nucleotide sequences. PCR mutagenesis procedures make it possible to modify and engineer any target DNA with ease and very high efficiency. This includes the ability to restructure genes by, for example, domain swapping experiments. In this chapter we will cover the basic principles of PCR-based mutagenesis for deletion and insertion of sequences, as well as introducing point mutations and approaches for recombination and the incorporation of random mutations.
