Polymerase Chain Reaction (PCR) is used to clone segments of DNA present in samples for identification of individuals, species and traits for a variety of different purposes. PCR was developed in the 1980's and has been used in forensics, for detection of genes linked to disease, to create phylogenies of species and to identify new species. Only a very small amount of DNA is needed for this reaction, which can produce billions of copies in a matter of hours1. PCR consists of three major steps:
1. Denaturation: DNA is heated to break down bonds holding it together, and becomes single stranded at 95ºC.
2. Annealing: Primers are short strands of DNA that are complementary to the sequence of interest in the sample. During this step, the primers bind to the DNA.
3. Primer extension: A protein complex called DNA polymerase extends the primer from the ends in the 5’ to the 3’ direction, lengthening the DNA. A common DNA polymerase used for this step is derived from the organism Thermus aquaticus, a thermotolerant bacterium2.
These steps are repeated to generate many strands of the DNA segment of interest. Lastly, gel electrophoresis can be used to detect quantities of the cloned segments3. In studies of invasive species, PCR is used to amplify sequences to determine genetic relationships and diversity between founding and invasive populations.
Figure 1. PCR steps2
1McClean, P. 1997. Polymerase Chain Reaction (or PCR). North Dakota State University. Accessed 21 Mar. 2015. Available from: http://www.ndsu.edu/pubweb/~mcclean/plsc431/cloning/clone9.htm.
2Davidson College. 2000. The Polymerase Chain Reaction. Accessed 23 Mar. 2015. Available from: http://www.bio.davidson.edu/Courses/Molbio/MolStudents/01jerusin/pcr.html.
3South Dakota State University. 2002. Polymerase Chain Reaction. Accessed 23 Mar. 2015. Available from: http://www.sci.sdsu.edu/~smaloy/MicrobialGenetics/topics/in-vitro-genetics/PCR.html