Since fingerprinting came into use by detectives and police labs, many people believed its uniqueness can demonstrate guilt or innocence of a suspect.  However, did you ever think that a criminal can alter their fingertips by surgery?  It is a possibility due to improvement of medicine nowadays.  So what is next?  In 1984, forensic procedures took a huge step forward with the introduction of DNA Fingerprinting.  DNA, known as the blueprint of all life form, contains unique genetic information that is specifically present only in its own species.  Since DNA is extracted from any cell, tissue, and organ of a person, it cannot be altered by any known technique.  That is the beauty of DNA Fingerprinting over conventional fingerprinting methods.  Since its invention, DNA Fingerprinting has gained worldwide recognition and is being used not only in biological evidence by FBI and police, but diagnosis of inherited disorders, developing cures of inherited disorders and personal identification. 

The basic building blocks of DNA are the nucleotides.  It is made up of deoxyribose sugar, a phosphate group, and one of four nitrogen bases: Adenine, Cytosine, Guanine and Thymine.  These bases combine in very specific ways.  Adenine pairs only with thymine, and guanine pairs only with cytosine. The information contained in DNA is determined by the sequence of base pair along the sugar phosphate backbone.  Different DNA sequences are what differentiate living organisms or characteristics because they provide the instructions used to build amino acids and link them together into protein.

In order to visualize DNA sequence with simple laboratory technique, DNA electrophoresis is used.  However, DNA Electrophoresis only works then there are enough number of DNA sequence regions.  This is where PCR (Polymerase Chain Reaction) comes in.  PCR was developed and still owned by Roche Molecular Systems, Inc. and F. Hoffmann-La Roche Ltd.   It is like xeroxing certain DNA regions.  Polymerases are enzymes that are present in all living organism.  Their roles are to copy genetic material, proofread, and correct the copies of DNA.  To ease your understanding, you can imagine DNA replication during the S phase of the cell cycle. After an enzyme attaches to the DNA, the DNA double helix is unwound into two single strands, a molecule of a DNA polymerase binds to one strand of the DNA.  Then, it begins to move along the strand, using it as a template for synthesizing a leading strand of nucleotides and as the strand is copied, the double helix closes once again.  The PCR process mimics this process.  It require a piece of original DNA to be copied, two different primer molecules to bracket the unwound DNA, individual nucleotides to be used as building blocks, buffer solution, and Taq DNA Polymerase.  Two different primers are used because one is complementary to one DNA strand at the beginning of the target region and a second primer is complementary to the other strand at the end of the target region. 

Under certain conditions, the region of DNA will amplify to very large quantity in short period of time.  The PCR mixture follows three repeated steps: denaturation, annealing the primer, and replicating the DNA. 

·        The original piece of DNA is first denatured at 94-96^oC.  The double stranded helix is separated into single strands.

·        Primer is binding to each stranded DNA at a lower temperature, around 50-65^oC.

·        Temperature should be raised back to 72^oC for DNA replication.

You may also view PCR animation at Background/Theory page to enhance your understanding.  As amplification proceeds, the DNA sequence doubles each cycle.  Since PCR almost depends on temperature change, usage of a thermocycler is strongly recommended.  Three water bathes of 94-96^oC, 50-65 ^oC, 72 ^oC are required prior to PCR experiment.  For 25-30 PCR process, students have to move a test tube back and forth 75-90 times.  It can be a very tedious job, and surely, it can induce some human error.  Therefore, using water baths are not suggested, yet it is still doable under a tight budget. 

Following site has good insights of PCR Animation.

·          Dolan DNA Learning Center

·          The National Health Museum

·          The PCR article by Tabitha M. Powledge

Once enough DNA sample has been amplified, DNA electrophoresis can be done.  Electrophoresis is the process of moving molecules with electric currents.  Electric charge is transferred through the agarose gel.  DNA fragments process a slightly negative charge.  Similar to a magnet, opposite poles attract. DNA fragments are repelled by the negative electrode and attracted to the positive electrode.  Thus, DNA fingerprinting is nothing but the trace left by certain DNA molecules.   Size and weight of each DNA fragments make distinct trace marks which is the beauty of DNA fingerprinting.

After electrophoresis, methylene blue staining is conventionally used to stain the gel. However, in order to increase visibility, special stains such as CarolinaBLU™ stain or QUIKView DNA Stain are acceptable to use.  Increased sensitivity, reduced background, and reduced staining time surely improve DNA electrophoresis process.