Applications of PCR - Research Applications

Research Applications

PCR has been applied to many areas of research in molecular genetics:

• PCR allows rapid production of short pieces of DNA, even when nothing more than the sequence of the two primers is known. This ability of PCR augments many methods, such as generating hybridization probes for Southern or northern blot hybridization. PCR supplies these techniques with large amounts of pure DNA, sometimes as a single strand, enabling analysis even from very small amounts of starting material.

• The task of DNA sequencing can also be assisted by PCR. Known segments of DNA can easily be produced from a patient with a genetic disease mutation. Modifications to the amplification technique can extract segments from a completely unknown genome, or can generate just a single strand of an area of interest.

• PCR has numerous applications to the more traditional process of DNA cloning. It can extract segments for insertion into a vector from a larger genome, which may be only available in small quantities. Using a single set of 'vector primers', it can also analyze or extract fragments that have already been inserted into vectors. Some alterations to the PCR protocol can generate mutations (general or site-directed) of an inserted fragment.

• Sequence-tagged sites is a process where PCR is used as an indicator that a particular segment of a genome is present in a particular clone. The Human Genome Project found this application vital to mapping the cosmid clones they were sequencing, and to coordinating the results from different laboratories.

• An exciting application of PCR is the phylogenic analysis of DNA from ancient sources, such as that found in the recovered bones of Neanderthals, or from frozen tissues of Mammoths. In some cases the highly degraded DNA from these sources might be reassembled during the early stages of amplification.

• A common application of PCR is the study of patterns of gene expression. Tissues (or even individual cells) can be analyzed at different stages to see which genes have become active, or which have been switched off. This application can also use Q-PCR to quantitate the actual levels of expression

• The ability of PCR to simultaneously amplify several loci from individual sperm has greatly enhanced the more traditional task of genetic mapping by studying chromosomal crossovers after meiosis. Rare crossover events between very close loci have been directly observed by analyzing thousands of individual sperms. Similarly, unusual deletions, insertions, translocations, or inversions can be analyzed, all without having to wait (or pay for) the long and laborious processes of fertilization, embryogenesis, etc.