The Polymerase Chain Reaction (PCR) procedure is a major laboratory technique used in the Human Genome Project for amplification of sequence fragments. One of the difficulties encountered when using PCR is that the product can be contaminated with the DNA of non-genomic sequences, most notably mitochondrial DNA. PCR is initiated by using small sequences, usually 20 or fewer bases, that act as primers for the reaction. This work is an attempt to recognize primers to avoid, in the sense that they will cause mitochondrial DNA to amplify and contaminate the genomic DNA product. Computer programs were written that, for a given primer, first find all locations of them in human mitochondrial DNA, and then compute whether PCR amplification can be expected, based on combinations of primer locations that are known to cause amplification. One of the programs being written analyzes the frequency of occurrence of all possible 3- to 10-mers of the four bases -- A, C, G, T == in given sequences. This program will also enable comparison of all known mitochondrial sequences. Data are being analyzed which indicate different patterns of sequence usage based upon the species of origin of the mitochondrial DNA. These analyses may yield valuable information on the mitochondria of the different organisms, as well as information pertinent to the regulation of mitochondrial DNA function and structure.
