Gel Electrophoresis

Question 1

What is electrophoresis?

Answer 1

This is the movement of molecules by an electric current

Question 2

What charge does DNA have and why?

Answer 2

DNA is negatively charged. This is due to its phosphate groupsb

Question 3

Towards what pole does DNA move to?

Answer 3

The positive pole (anode)

Question 4

How is DNA electrophoresis carried out?

Answer 4

DNA is applied to a gel matrix, and then e,extricate current pulls the DNA through that gel matrix which slows its progress towards the positive side

Question 5

What is therefore the relationship between DNA speed and its size ?

Answer 5

DNA trace,s at speeds inversely related to its size. Big DNA molecules go slower while smaller ones GI faster and migrate further in the gel

Question 6

Why are gels used for electrophoresis?

Answer 6

Gels form a porous matrix that the DNA can squeeze through

Question 7

What are the two most common gels used in molecular biology?

Answer 7

Agarose

Polyacrylamide

Question 8

What us agarose gel?

Answer 8

It is a polymer from seaweed and a component of the agar used for bacterial culture

Question 9

Agarose is used for what type of DNA molecules?

Answer 9

Larger DNA molecules of 50bp to 50000bp

Question 10

What does the concentration of agarose determine?

Answer 10

The sizes of the spaces in the gel

Question 11

What kind of pores do high concentrations of agarose make?

Answer 11

Smaller spores

Question 12

Low concentration of agarose results in what type of pores?

Answer 12

High pores

Question 13

In what concentrations is agarose used?

Answer 13

0.5-5%

Question 14

High percentages of agarose are used for

Answer 14

Smaller DNA molecules

Question 15

Low percentages of agarose are used to resolve

Answer 15

Larger DNA molecules

Question 16

Polyacrylamide gel electrophoresis is used to resolve what type of DNA fragments?

Answer 16

Very small DNA fragments and ss DNA down to 1bp

Question 17

PAGE is often used for what type of analysis?

Answer 17

Sequencing and mutation analysis

Question 18

In what direction is PAGE typically run? Why?

Answer 18

Vertically, it’s a potent neurotoxin

Question 19

What’s the first step when making an agarose gel?

Answer 19

Making the electrophoresis buffer

Question 20

What is the use of buffer in electrophoresis?

Answer 20

The buffer carries the current and protects the samples during electrophoresis

Question 21

What are the two most common types of buffer? And how how do they differ?

Answer 21

Tris acetate and Tris borate. Tris acetate moves faster but with a smaller buffering capacity while Tris borate moves slower with a better buffering capacity

Question 22

What does the loading dye added to the DNA sample do?

Answer 22

Gives color to DNA for easier visualization
Makes DNA denser than water so it stinks to bottom of well
Has tracking dyes that separate during electrophoresis to indicate progress of electrophoresis

Question 23

What is a molecular weight standard?

Answer 23

It is a concentrated control stock of DNA fragments of known size