Biology 160
Name and Lab Section________________
Forensic DNA: DNA Profiling
Introduction:
In the next three labs you will become crime scene detectives and help determine which suspect
was at the scene of the crime. To do this you will compare the DNA “fingerprints” of blood cells that
was found at the crime scene and compare that to the DNA “fingerprints” of several suspects.
Objectives:
Be able to WOW your friends by having the ability to describe in detail how DNA profiling
works. You will also have experience making a DNA “fingerprint”.
To do that you will
a) Be able to explain what is meant by a “DNA profiling or a DNA fingerprint”
b) Understand what is meant in molecular biology by the phrase “cut and run”
c) Understand the use of Restriction Enzymes in cutting up DNA
d) Understand how Gel Electrophoresis separates different sizes of DNA
e) Be able to quantify the size of DNA using DNA markers
f) Understand what PCR stands for and when it is used
DNA Profiling or Fingerprinting:
DNA profiling is based on the fact that Individuals have different sequences of DNA nucleotides. This
allows a person to be identified by their DNA. This is a several step process that involves cutting the
DNA with restriction enzymes, separating the DNA using gel electrophoresis and staining the DNA
fragments.
Restriction Enzymes:
Restriction enzymes, or endonucleases, are enzymes that cut DNA at specific sequences of nucleotides.
The specific sites they cut at are called restriction sites. Since different individuals can have different
sequences of nucleotides they will not all have the same restriction sites. This is referred to restriction
fragment length polymorphisms (RFLP). Also in the human genome there are short sequences of DNA
that repeat called short tandem repeats (STR). Different individuals have different numbers of these
repeated sequences. Therefore the distance between two restriction cut sites can be different for
different people. This (STR analysis) can also be used to identify individuals.
Agarose Gel Electrophoresis:
Agarose gel electrophoresis separates DNA fragments by size. DNA is loaded into an agarose gel that
is attached to a power supply. When a current is passed between electrodes in the apparatus, the DNA
will move towards the positive end of the gel since the phosphate groups on the backbone of DNA
have a negative charge. The smaller fragments of DNA move quicker through the gel and the
fragments of DNA become separated by size. To see the DNA, a stain is added and bands of DNA
become visible.
Polymerase Chain Reaction:
Polymerase Chain Reaction (PCR) is a way to quickly make many copies of DNA. In many instances,
including crime scenes, before DNA can be “cut and run”, more needs to be made. We have enough
DNA for our analysis so we will NOT need to perform PCR.
Please see your textbook, pages 242 through 247 for more detail on these techniques.
Overview:
Day 1:
During the first laboratory session you will practice using pipettors. After you have practiced with the
pipettors you will use restriction enzymes to cut the DNA of the suspects and the DNA found at the
crime scene. You will also be introduced to the gel apparatus that will separate the pieces of DNA so
that in our next lab you will be able to quickly set-up the apparatus.
Day 2:
During this laboratory session you will prepare the agarose gel and the electrophoresis apparatus. You
will then load the gel with the DNA that you prepared (cut) in the previous lab. After the DNA has
separated you will remove the gel from the electrophoresis apparatus and put it in a tray with stain
which will make the DNA visible. While the DNA is “running” you will answer questions about the
procedure.
Day 3:
You will look at the pattern of DNA fragments and determine which suspect was most likely at the
crime scene. You will also determine the size of the DNA fragments by making a “standard curve.”
The specific laboratory procedures will be given to you in the lab.