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Why did you add Laemmli sample buffer to your fish samples?
What was the purpose of heating the samples?
Why do SDS-coated proteins move when placed in an electric field?
What is the purpose of the actin & myosin standards and the Precision Plus Protein Kaleidoscope
prestained standard?
Which proteins will migrate farthest? Why?
What is the purpose of the Coomassie blue stain?
How many individual protein subunits make up the quaternary structure of one biologically active
myosin protein? What are these proteins? What are their approximate molecular masses?
What effect does heating the sample have on the extracted material?
Why is it important to denature proteins before electrophoresis?
Which two fish have the most similar protein profiles?
Which two fish have the least similar protein profiles?
Why are proteins treated with ionic detergent (SDS), reducing agents (DTT), and heat before SDSPAGE (gel electrophoresis)?
What is the purpose of using experimental controls? What purpose do the actin & myosin standards
serve? The prestained standards?
Why is it important for the gel to be in complete contact with the membrane without any air bubbles?
Why do proteins migrate from the gel to the membrane?
Why are proteins blotted from the polyacrylamide gel to a membrane?
Can you think of a way to determine if the transfer of an unstained gel has been successful?
Describe how a specific protein can be identified from a mixture of proteins.
What information does the western blot provide for each sample?
The molecular mass of myosin light chain 1 is approximately 22 kD, myosin heavy chain is 200 kD
and actin is 42 kD. Which proteins will migrate fastest through the gel? Why?
Draw a gel and mark the relative positions of myosin light chain 1, myosin heavy chain, and actin
from the actin and myosin standard after electrophoresis.
Are myosin proteins the same or different sizes across species? How are protein sizes calculated?
How can this information be used to explain structural (and perhaps evolutionary) differences in
animal species?
Explain why the secondary antibody is used.
Give an explanation for why you think the protein profiles of some fish species share more bands
than other fish species.
What are 5 proteins found in muscle? What do they do?
What is the difference between the primary and quaternary structures of proteins?
Draw, label, and describe the main quaternary structure of myosin, including all protein subunits.
Why has the structure of actin and myosin been conserved over millions of years?
How do variations in organisms occur in nature, and why? How does this contribute to biodiversity?
How might variations in proteins between species be used to determine their evolutionary
relationships?
How can diverse species share so much common DNA sequence?
Can one gene encode more than one protein? How can two different proteins derived from the same
gene have different sizes and have different functions?