Wright 1 - Electrophoresis

Question 1

What is the positive electrode called?

Answer 1

The anode

Question 2

What is the negative electrode called?

Answer 2

cathode

Question 3

DNA moves in what direction in electrophoresis? Which electrode to which electrode?

Answer 3

From the cathode (-) to anode (+)

Question 4

What is agarose?

Answer 4

An uncharged polysaccharide purified from agar of the seaweed Agar ager. To prepare it agar is harvested in the sol state and heated to make the initial gel and then aged to make the final gel structure

Question 5

What type of filter must be placed below a gel when UV light is shined onto it from below?

Answer 5

Transiluminator filter

Question 6

How do linear DNA molecules migrate through gel? (eg at what rate?)

Answer 6

At a rate that is inversely proportional to the log of their molecular mass or base pairs

Question 7

What three parameters influence the rate of migration of DNA molecules during gel-electrophoresis?

Answer 7

1. Percentage of agarose in gel-matrix
2. Topology (conformation) of the DNA molecule: linear, relaxed circular, or supercoiled
3. The voltage applied (<5 volts/cm)

Question 8

How does agarose concentration effect DNA molecule migration?

Answer 8

• Higher concentration decreases average pore size

- Smaller pore size cause less DNA migration and effects the separation range of linear DNA molecules

Question 9

How can separation range of DNA be changed in electrophoresis?

Answer 9

By altering agarose concentration

Question 10

What are the three topologies that DNA molecules can take?

Answer 10

• Linear
• Relaxed circle
• Supercoiled circle

Question 11

What is the most common type of supercoil in DNA?

Answer 11

Negatively supercoiled

Question 12

How can positively supercoiled DNA be observed?

Answer 12

in vitro

Question 13

Rank the topologies of DNA from least distance migrated to most distance migrated

Answer 13

Least distance

1. Relaxed circular
2. Linear
3. Supercoiled

Question 14

Does %GC content influence rate of migration of DNA molecules during agarose gel electrophoresis?

Answer 14

no

Question 15

What do restriction endonucleases do for bacteria and some yeast?

Answer 15

Prevent infection by DNA viruses

Question 16

How can you name a restriction endonuclease?

Answer 16

1. First letter of genus name (eg. E)
2. Couples letters of species name (eg. co)
3. First letter of strain name (eg. R)
4. Order of indentification (eg. I, first identified)

Altogether that makes EcoRI

Question 17

Why do geneticists care about mapping recognition sites for restriction endonucleases?

Answer 17

It is often the first step in the characterization of an unknown DNA molecule, leading to further manipulations. Eg. cloning and DNA sequencing