Thermostable Ligase-Mediated Incorporation of Mutagenic Oligonucleotides During PCR Amplification

Abstract

PCR amplification has become an extremely powerful and universally used technique for cloning and manipulating DNA segments. It is especially useful for the ability to alter the terminal sequences of an amplified product simply by using primers containing the desired changes. However, the inability to alter easily regions of a product between the amplification primers has limited the use of PCR for more general mutagenesis. The protocol presented in this chapter employs a sample method that removes this limitation.