ll|llllManual Supplement Multiplex PCR: Advantages, Development, and Applications

Institute for Molecular Genetics, Baylor College of Medicine, Houston, Texas 77030 Ever since it was shown that PCR could simultaneously amplify multiple loci in the human dystrophin gene, (1) multiplex PCR has been firmly established as a general technique. A short list of multiplex PCR applications now includes pathogen identification, gender screening, linkage analysis, forensic studies, template quantitation, and genetic disease diagnosis. Multiplex PCR can be a two-amplicon system or it can amplify 13 or more separate regions of DNA. It may be the end point of analysis, or preliminary to further analyses such as sequencing or hybridization. The steps for developing a multiplex PCR and the benefits of having multiple fragments amplified simultaneously, however, are similar in each system. This review focuses on multiplex systems in which each primer pair targets a single locus, unlike RAPD (2) and alumorph {3) PCR reactions, which amplify multiple loci with a single primer or primer set.