Biotechnology: Gel Electrophoresis

Question 1

Purpose of Gel Electrophoresis

Answer 1

A technique that can separate large, charged molecules according to size and charge, so they can be visualised and identified

Question 2

Uses of Gel Electrophoresis

Answer 2

1. Forensics

2. Paternity/relatedness

Question 3

Direction of Electrical Charge

Answer 3

DNA molecules have an overall negative charge due to the phosphate of the nucleotide, so they are repelled from the negative electrode and attracted by the positive electrode

Question 4

Components

Answer 4

• Agarose gel
• DNA ladder
• Buffer solution
• Positive and negative electrodes

Question 5

Why is a buffer solution used?

Answer 5

It allows a current to be carried through the sample and maintain the gel at a stable pH

Question 6

DNA ladders (size markers)

Answer 6

Pieces of DNA of a known number of base pairs used to determine the size of unknown DNA fragments

Question 7

Purpose of agarose gel

Answer 7

It allows an electrical current to run through it and acts as a sieve, with smaller molecules moving through it faster and larger ones moving slower

Question 8

Step 1: Prepare Gel

Answer 8

Agarose gel is melted and poured into a flat mould to cool

Question 9

Step 2: Add Samples

Answer 9

Wells are created in the gel as it sets and the DNA samples and a DNA ladder are pipetted into separate wells

Question 10

Step 3: Buffer + Electrodes

Answer 10

The gel is placed in a tray with buffer solution and positive and negative electrodes are attached at either end

Question 11

Step 4: Direction of Charge

Answer 11

When a current runs, the fragments are repelled by the negative electrode and more toward the positive

Question 12

Step 5: Movement of the Bands

Answer 12

The gel acts as a large sieve and smaller strands of DNA move through it faster, while larger strands move slower, separating the DNA according to size

Question 13

Step 6: Visualising

Answer 13

To view the separated DNA fragments ethidium bromide or another florescent DNA-binding dye is added to the agarose gel before it sets

Question 14

DNA Profiling

Answer 14

A technique used to identify an individual by comparing an unknown sample of DNA with known DNA profiles

Question 15

Short Tandem Repeats (STR’s)

Answer 15

Short, repeating stretches of non-coding DNA around 2-5 base pairs in length, used in gel electrophoresis

Question 16

Difference between DNA Profiling and Sequencing

Answer 16

• Profiling identifies individual by looking at unique band patterns
• Sequencing determines sequence of nucleotides
• Profiling is faster and more visual
• Profiling allows you to test more samples