Electrophoresis Exam
60 Points
Date________________
Name_____________________________
(4 points each, 24 points total) MULTIPLE CHOICE
__D__ 1) All of the following are reasons to use TRIS during an electrophoresis except
A) To stabilize the pH
B) To conduct electricity
C) To maintain the shape of the molecules
D) To stain the molecules
__D__ 2) Which stain causes DNA to glow orange when exposed to UV light?
A) Agrose
B) Polyacrylamide
C) Methylene blue
D) Ethidium bromide
__C__ 3) A circular plasmid was cut with a restriction enzyme and three bands were
visualized after electrophoresis. How many cuts were made in the plasmid?
A) 1
B) 2
C) 3
D) 4
__A__ 4) What is Agar?
A) Solid media on which bacteria grow
B) Liquid media for bacterial growth
C) Region on a bacterial operon
D) Matrix used for vertical get electrophoresis
“This workforce solution was funded by a grant awarded under the President’s Community-Based Job Training Grants as
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__C__ 5) How can colorless proteins be monitored during electrophoresis?
A) Coomassie blue stain
B) Silver stain
C) PAGE loading dye
D) Ethidium bromide
__C__ 6) What is the purpose for SDS during electrophoresis?
A) Stain the proteins for visualization
B) Use as a loading dye
C) Denature the proteins and give them a negative charge
D) Conduct electricity
7. (8 Points) List and explain the factors that DNA migration in Agrose depend on.
1. Agarose Concentration
Using different concentrations gels resolve different sizes of DNA fragments
– Higher concentrations of agarose for small DNAs
– Low agarose concentrations allow resolution of larger DNAs
2. Voltage
As voltage applied to a gel is increased, larger fragments migrate proportionally faster that
small fragments
3. Electrophoresis Buffer
_DNA fragments will migrate at somewhat different rates in these two buffers due to
differences in ionic strength
_Buffers not only establish a pH, but provide ions to support conductivity
4. Effects of Ethidium Bromide
_Binding of ethidium bromide to DNA alters its mass and rigidity, and therefore its mobility
8. (4 Points) Explain what is meant by amphoteric compounds? How does that affect the
compound’s mobility in gel electrophoresis? Be specific.
– Their net charge therefore is determined by pH of medium in which they
are suspended
• In a solution with a pH above its isoelectric point, a protein has a
net negative charge and migrates towards anode in an electrical
field
• Below its isoelectric point, protein is positively charged and
migrates towards cathode
“This workforce solution was funded by a grant awarded under the President’s Community-Based Job Training Grants as
implemented by the U.S. Department of Labor’s Employment and Training Administration. The solution was created by the grantee
and does not necessarily reflect the official position of the U.S. Department of Labor. The Department of Labor makes no guarantees,
warranties, or assurances of any kind, express or implied, with respect to such information, including any information on linked sites
and including, but not limited to, accuracy of the information or its completeness, timeliness, usefulness, adequacy, continued
availability, or ownership. This solution is copyrighted by the institution that created it. Internal use by an organization and/or
personal use by an individual for non-commercial purposes is permissible. All other uses require the prior authorization of the
copyright owner.”
9. (4 Points) What is SDS? Why does it get added to proteins in electrophoretic
separations?
– sodium dodecyl sulfate (SDS)
• Denatures proteins into polypeptide strands
• Gives each polypeptide strand an overall negative charge
10. (20 Points) Fill in the blank places at the table below:
Horizontal or vertical?
Why are they poured
horizontal or vertical?
Separates large
molecules or small
ones?
Lets you tell small
differences between
sizes of bands?
Can be reheated and
poured?
Powder is toxic?
Solid is toxic
Agarose
Horizontal
Because you can. It’s easier to
pour a horizontal gel than it is
to pour a vertical gel.
Both, but better for large
molecules
Polyacrylamide
Vertical
Need glass plates for
structure- very thin gels that
need support.
Air speeds up
polymerization. If the gel
was poured open to air, it
would not solidify evenly.
Both, but better for small
molecules
No
Yes
Yes
No
No
No
Yes
No
“This workforce solution was funded by a grant awarded under the President’s Community-Based Job Training Grants as
implemented by the U.S. Department of Labor’s Employment and Training Administration. The solution was created by the grantee
and does not necessarily reflect the official position of the U.S. Department of Labor. The Department of Labor makes no guarantees,
warranties, or assurances of any kind, express or implied, with respect to such information, including any information on linked sites
and including, but not limited to, accuracy of the information or its completeness, timeliness, usefulness, adequacy, continued
availability, or ownership. This solution is copyrighted by the institution that created it. Internal use by an organization and/or
personal use by an individual for non-commercial purposes is permissible. All other uses require the prior authorization of the
copyright owner.”
Type of molecules run
in this type of gel
Big gaps or small gaps
in gel?
Expense?
Mostly DNA
DNA or proteins
Bigger gaps than with
polyacrylamide
Agarose is about $1 a gramconsidered expensive.
Smaller gaps
Gels usually bought prepoured. Cost $5-7 per gel.
“This workforce solution was funded by a grant awarded under the President’s Community-Based Job Training Grants as
implemented by the U.S. Department of Labor’s Employment and Training Administration. The solution was created by the grantee
and does not necessarily reflect the official position of the U.S. Department of Labor. The Department of Labor makes no guarantees,
warranties, or assurances of any kind, express or implied, with respect to such information, including any information on linked sites
and including, but not limited to, accuracy of the information or its completeness, timeliness, usefulness, adequacy, continued
availability, or ownership. This solution is copyrighted by the institution that created it. Internal use by an organization and/or
personal use by an individual for non-commercial purposes is permissible. All other uses require the prior authorization of the
copyright owner.”