Polymerase Chain Reaction
Author: Rodney E. Shackelford, D.O., Ph.D. (see Reviewers page)
Revised: 8 July 2010, last major update July 2010
Copyright: (c) 2008-2010, PathologyOutlines.com, Inc.
● In multiplex PCR, several discreet sequences are amplified simultaneously in the same reaction using multiple primer pairs
● The technique is used to verify that an amplifiable sequence is present in a specific sample, or to amplify multiple sequences within one reaction
● Usually the sequence being searched for is co-amplified with a known “housekeeping gene”, which is always present and easily amplified
● If only the housekeeping gene is amplified, then the other sequence is absent from the original sample
● If neither sequence is amplified, then a PCR inhibitor is likely present
● Up to 20 different reactions can be run simultaneously, thus lowering the amount of sample used, reducing the reagents consumed, and collecting far more information per reaction, while simplifying data analyses
● This technique is widely used in forensics, tissue identification, detection of different species of viral nucleic acids within cerebrospinal fluid and transplantation engraftment studies, where it can use single nucleotide polymorphisms and other variation in DNA sequences to identify tissue sample origins
● The main disadvantages are the difficulty, sometimes extreme, in optimizing the PCR reaction so that each sequence amplification is roughly as efficient as the other reaction within the same reaction chamber, and designing primers that will not interact with each other (i.e., “primer dimmers), resulting in non-specific amplifications
End of Molecular Pathology > Polymerase Chain Reaction > Multiplex PCR
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