Molecular Genetics ❮ 129similar sticky ends. The DNA can be attached to the plasmid, creating a vector that can be
used to transport DNA.Gel Electrophoresis
This technique is used to separate and examine DNA fragments. The DNA is cut with our
new friends, the restriction enzymes, and then separated by electrophoresis. The pieces of
DNA are separated on the basis of size with the help of an electric charge. DNA is added
to the wells at the negative end of the gel. When the electric current is turned on, the migra-
tion begins. Smaller pieces travel farther along the gel, and larger pieces do not travel as far.
The bigger you are, the harder it is to move. This technique can be used to sequence DNA
and determine the order in which the nucleotides appear. It can be used in a procedure
known as Southern blotting (after Edwin M. Southern, a British biologist) to determine
if a particular sequence of nucleotides is present in a sample of DNA. Electrophoresis is
used in forensics to match DNA found at the crime scene with DNA of suspects. This
requires the use of pieces of DNA called restriction fragment length polymorphisms(RFLPs).
DNA is specific to each individual, and when it is mixed with restriction enzymes, differ-
ent combinations of RFLPs will be obtained from person to person. Electrophoresis sepa-
rates DNA samples from the suspect and whatever sample is found at the scene of the
crime. The two are compared, and if the RFLPs match, there is a high degree of certainty
that the DNA sample came from the suspect. In Figure 11.9, if well A is the DNA from
the crime scene, then well C is the DNA of the guilty party.Figure 11.9 A sample gel electrophoresis.ABCD
Cloning
Sometimes it is desirable to obtain large quantities of a gene of interest, such as insulin for
the treatment of diabetes. The process of cloning involves many of the steps we just men-
tioned. Plasmids used for cloning often contain two important genes—one that provides
resistance to an antibiotic, and one that gives the bacteria the ability to metabolize some
sugar. In this case, we will use a galactose hydrolyzing gene and a gene for ampicillin resis-
tance. The plasmid and DNA of interest are both cut with the same restriction enzyme.
The restriction site for this enzyme is right in the middle of the galactose gene of the plas-
mid. When the sticky ends are created, the DNA of interest and the plasmid molecules are
mixed and join together. Not every combination made here is what the scientist is looking
for. The recombinant plasmids produced are transformed into bacterial cells. This is where
the two specific genes for the plasmid come into play. The transformed cells are allowed to
reproduce and are placed on a medium containing ampicillin. Cells that have taken in theKEY IDEASteve (12th grade):
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