PCR-based strategy of constructing chimeric DNA molecules

摘要

The invention relates to methods for the use of the polymerase chain reaction to amplify a segment of a cloned gene of interest in such a way as to allow a simplified introduction of alterations such as deletions, insertions, repetitions (both direct and inverted) and substitutions into the cloned gene in a specific and precise manner. The unique, amplified segment of the cloned gene so amplified is a common substrate for each of the different approaches to introducing the various alterations into the gene. Choice of the primer sites within the amplified segment coupled with choice of the orientation of the molecule once ligated to itself results in the various resulting embodiments of the invention.