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DNA Fingerprinting

How DNA Fingerprinting Works

How to produce a genetic fingerprint:

The first step to making a genetic fingerprint requires getting a sample of DNA.  This sample can come from blood, semen, hair or saliva, and may be an extremely small sample.  The root from a single strand of hair is enough for researchers to work with.  This sample contains white blood cells which are broken open using detergent, and all the useable DNA is separated from the extra cellular material.  Next the restriction enzymes are used to cut the DNA into smaller pieces.  Restriction enzymes work by cutting the DNA at a specific sequence, which produces either blunt ends or sticky ends, and results in many fragments of different lengths. These fragments are called restriction fragments length polymorphisms, or RFLPs.

These RFLPs are then put into an agarose gel.  Using gel electrophoresis, the fragments are sorted according to size.  When the current of the electric field is turned on, the negative RFLPs will start to move across the gel towards the positive end.  The smaller fragments move farther across the gel than the larger ones.  Also, alkali is responsible for causing the hydrogen bonds to break, and the DNA to become single-stranded.  When the DNA becomes single-stranded, it causes nucleotides to become free, and they will later be used to pair up with probes.  The gel is then covered by a piece of nylon and thin paper towels, which are used to absorb moisture from the gel.  The DNA fragments get gently transferred from the gel to the surface of the nylon.  This process is called blotting.

 

Finally, radioactive probes get washed over the nylon surface.  These probes will join to any DNA fragments that share the same composition.  The final step to making a genetic fingerprint is to place a photographic film on top of the nylon surface.  The probes leave marks on the film wherever they attached to the RFLPs.  Dark bands will then show up when the film is developed, which marks the length of the RFLPs that were hybridized.  Researchers are then able to read the fingerprint and match it to others.  They do this by placing the xray on a light background, and comparing the RFLP lengths in the DNA from the crime scene, to the DNA of the suspect. 

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This picture shows DNA being sorted by an electric field.  When the current is turned on, it attracts the shortest and lightest particles the farthest.  Click to view site.

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When fingerprinting was a fairly new process, during 1984-1990, it could take as long as four to six weeks of lab work to complete and compare DNA fingerprinting evidence.  Jeffreys was responsible for coming up with the original DNA fingerprinting technique and in November of 1991, he also was responsible for creating a better test that could obtain results in as little as two days.
 
Polymerase Chain Reaction (PCR)
PCR can be used in DNA fingerprinting as a way to make numerous copies of isolated DNA.  The process can selectively amplify a single copy of a desired sequence. 
First Cycle:
-Dna molecules are melted by raising the temperature to 95 degrees Celcius.
-Strands separate, temperature is lowered to 60 degrees Celcius.
-Each primer binds specifically to the 3 prime end of the target sequence on the appropriate strands of DNA.
-Primers direct taq polymerase to synthesize complementary nucleotides.
-Only DNA containing target sequence are copied by taq polymerase. 
-At the end of cycle 1, both strands have been copied to form 2 partially double-stranded molecules.
-These steps are repeated during the next cycles.  After two cycles, four partially double-stranded molecules are produced, all containing the target sequence.
-After many cycles, millions of copies of the DNA can be produced. 
-This is useful in DNA fingerprinting, as researchers can do many tests on the same DNA sample.  

An Overview of How DNA Fingerprinting Works

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This diagram shows the detailed process of creating a DNA fingerprint.  Click to view site.

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