The Polymerase Chain Reaction (PCR)

The Polymerase Chain Reaction is a technique used to amplify a particular fragment or sequence of DNA.

PCR occurs when two oligonucleotide primers anneal to a DNA template in a proximity and orientation which allows the DNA polymerase to synthesize the DNA sequence which lies between the two primers.

A PCR reaction is comprised of:

a double-stranded DNA molecule: this is the "template" which contains the sequence to be amplified.
primer(s): this is a single-stranded DNA molecule which can anneal (bind) to a DNA sequence in the template DNA which has the complementary sequence.
dNTPs: a mixture with equal amounts of dATP, dTTP, dGTP, and dCTP which are the nucleotide subunits which will be put together to form new DNA molecules in the PCR amplification procedure.
Taq DNA polymerase: the enzyme which synthesizes the new DNA molecules using the dNTPs.

The PCR reaction involves numerous repeated cycles of DNA synthesis. Each cycle uses the products of the previous cycle as a DNA templates. After 30 - 40 cycles, a single DNA template molecule can produce millions of DNA products containing the sequence found between the two primers. (To see how the amplification works, click here.)

How PCR can be used to distinquish between two different DNA sequences.

Each PCR primer anneals only to a sequence in the DNA template which is complementary to its own.

If primer #1 and primer #2 can anneal to Genome A, then PCR can occur and a DNA product will be synthesized in the PCR reaction. However, if Genome B differs from Genome A in the sequence to which primer #2 anneals to the DNA, then PCR cannot occur successfully using Genome B DNA as a template, and no DNA product is synthesized.

Thus, this pair of primers (#1 and #2) allows us to distinguish between Genome A and Genome B.

Genome A and Genome B can represent genomic DNA from two individuals in the same species or possibly from two different species. Certain portions of genomic DNA tend to very conserved (very little variation) while other portions tend to vary greatly among individuals within a species or among different species.

The trick in such PCR analysis is to:

find those sequences which have just enough variation to allow us to detect differences among the organisms that we are studying.
find the right PCR primers which will allow us to detect sequence differences.

RAPD Analysis is a type of PCR reaction which also allows us to detect differences in various genomes.