Choosing highly specific primers for the polymerase chain reaction using the octomer frequency disparity method:...

The eight-nucleotide sequence (octomer) at the 3' end of PCR primers is important to PCR specificity. We describe a correlation between the specificity of PCR primers used with human DNA and the frequency of the 3' octomer in a human database. We therefore applied a methodology (OFD) based on octomer frequency disparity to identify 16 PCR targets in the chromosome of the intracellular bacterium, Chlamydia trachomatis (Ct). In addition, the 16 sets of primers were tested with a standard procedure. All the primer pairs were highly specific for Ct and did not lead to non-specific amplification when used with human DNA. This work shows that the choice of specific PCR primers is possible using a method based on the statistical representativeness of octomers in genomes.