Gel Electrophoresis and DNA Fingerprinting

1
Gel Electrophoresis and DNA Fingerprinting
2
What is gel electrophoresis?

• Because DNA is too small to see with the naked
  eye or with a microscope, we need some way to
  visualize DNA
• Gel electrophoresis is a way to visualize DNA by
  cutting it into pieces, staining the pieces, and
  separating the pieces by size

3
How do we cut the DNA?

• A restriction enzyme can cut DNA when it
  recognizes a certain sequence in the DNA

4
Restriction Enzymes

• Restriction Enzymes recognize certain
  palindromic DNA sequences
• These enzymes are found naturally in bacteria as
  protection against foreign (viral) DNA
• When the enzymes cut, they leave either blunt or
  sticky ends

5
What kind of DNA gets cut?

• If you are trying to create a unique DNA
  fingerprint, you would cut intron DNA
• Introns are segments of DNA that dont code for a
  protein
• Intron sequences are highly variable from person
  to person whereas exon sequences are very similar
• Each person has unique numbers and locations of
  restriction enzyme cut sites in their intron
  sequences

6
How do the pieces of DNA separate in gel
electrophoresis?

• Pieces of DNA are loaded into a well in a piece
  of gel
• DNA has a negative charge due to its phosphate
  group
• DNA is attracted to a positive electrode at the
  end of the gel

7
Size Separation

• The gel is made up of dense fibers, somewhat like
  a dense forest
• As DNA moves through the gel, smaller pieces can
  weave through more quickly and larger pieces get
  left behind

8
Procedure Pouring the Gel

• The first step is to tape the ends of the tray
  and insert a comb to make the wells.
• Then heat the gel ingredients, cool the solution
  to 65 degrees, and pour into the tray
• When cool, remove tape, place tray in the
  chamber, pour buffer solution over the tray, and
  wiggle out the comb

9
Procedure Loading the DNA

• Using a micropipette, load DNA into each well
• Always use a clean tip on the end of the pipette
  for each sample
• Be careful not to poke down too deep in the gel
  and puncture the well
• Ideally the DNA is released in the middle of the
  well

10
Procedure Running the Gel

• The lid of the gel chamber is plugged in to a
  power supply
• The apparatus is switched on and the fragments of
  DNA will start to move through the gel

11
Procedure Analyzing Results

• After the fragments are allowed to spread through
  the gel, the gel is stained so that the DNA bands
  become visible
• The sizes of the fragments become apparent by the
  position of the bands on the gel

12
An Example of DNA Fingerprinting
13
DNA Fingerprinting

• So to use this procedure, you would take the
  evidence and suspect DNA, cut it with restriction
  enzymes, and then run it on a gel
• If the bands are the same between suspect and
  evidence, you can assume that the suspect is
  guilty

If lane A is evidence from saliva from a dog
bite, and B, C and D are suspect dogs, which dog
is guilty?
14
Summary

• 1- Cut DNA with restriction enzymes
• 2- Load the fragmented DNA onto the gel
• 3- Run the gel to separate DNA by size
• 4- The resulting pattern is unique to every
  person and is called a DNA Fingerprint