“WHO’S DNA WAS LEFT BEHIND?”
PRINCIPLES AND PRACTICE OF AGAROSE GEL ELECTROPHORESIS
PRELAB
1. What is gel electrophoresis used for in science?
2. What determines the separation of molecules during the process of electrophoresis?
3. DNA, RNA, and proteins are all ______________ and hence capable of being separated by
electrophoresis.
4. The powder that is used in making the gel belongs to which macromolecule group?
5. What is the purpose of mixing glycerol with the samples used in electrophoresis?
6. What is the purpose of using electricity in the electrophoresis apparatus?
7. Does DNA have a net negative or positive charge?
8. Since DNA has this charge, which electrode will it move toward: anode (+) or cathode (-)?
9. What 3 factors affect the mobility of the molecules (DNA) moving through the gel?
10. What is “DNA Fingerprinting” and what is it used for?
11. Name two human substances in which a forensic scientist might collect to be fingerprinted?
12. What are restriction enzymes and how are they used in electrophoresis?
13. What do the dyes in this experiment represent?
14. The ____________ _______________ serves as the individual “fingerprint”.
15. What is the objective of this laboratory investigation?
16. What is the independent variable? Dependent variable?
17. Due to this being an “introductory” experiment, dye will be used instead of a molecule like DNA.
How many different dyes will be loaded into the electrophoresis gel?
18. What sample (crime scene and/or suspect) does each dye (A-F) correspond to?
19. What volume of dye will be loaded into each well?
20. How might the dyes differ from one another (bottom of page 4 “Background Information”)?
21. Which electrode will the dyes move toward?
22. How long will it take to verify if the experiment is working properly? How will you know?
23. Write a hypothesis for this experiment.
Familiarize yourself with the following diagrams and explanations.
Below is an actual picture of the electrophoresis apparatus and a micropipette. Notice that the
apparatus has a black cord (cathode) and a red cord (anode) at opposite ends on the lid. The actual
apparatus is several centimeters deep where the gel and buffer will be placed.
Micropipettes are used to distribute extremely small quantities of a liquid. They measure in
microliters (µL). The ones you will be using are incremented from 20-200 µL. Remember that a
microliter is 1000th of a milliliter.
Below is a diagram showing an electrophoresis experiment that is ready to be “ran”. The gel has
been placed into the electrophoresis “chamber”, the samples have been “loaded” and the electricity
is ready to be turned on.
Notice the negative and positive electrodes. The
sample wells (“samples” in the diagram) are
placed at the negative end of the apparatus. Since
agarose electrophoresis is used to separate
nucleic acids, and since they have a negative
charge, they will migrate from the negative end to
the positive end (shown by the arrow). The wells
are indentions in the gel, which is a centimeter or
two thick. The samples, such as DNA will be
“loaded” into the wells using a micropipette.
Below is a diagram showing the finishing results of an electrophoresis experiment. The samples have
finished migrating and the electricity has been turned off.
1 2
3 4
5
Notice the “bands” in the gel. Each band is
conglomeration of molecules, DNA, that share
similar properties (size, charge, and shape). The
larger DNA molecules migrated a short distance
due to their inability to get through the pores of
the gel. The smaller molecules migrated farther
due to being small enough to fit through the pore
size of the gel. This gel would have been run for
about 90 minutes for this to occur.
Imagine that the wells were labeled 1-5 from left
to right. Well #1 would have had eight different
segments of DNA ranging from the largest to the
smallest segments in this experiment. Well #2
would have had three segments of relatively large
size and well #5 would have had one large
segment of DNA.