Gene Cloning and Transformation

Question 1

What is gene cloning?

Answer 1

Gene cloning (DNA cloning) is a system that allows DNA fragments/genes to be copied.

Question 2

What are the basic steps of gene cloning?

Answer 2

• Restriction enzyme cuts up foreign DNA that contains a gene that will be copied
• The restriction enzyme cuts up the DNA of a cloning vector (plasmid)
• Mix cut foreign DNA with cut plasmids
• Apply DNA ligase to join fragments together
• Mix plasmids with bacteria to allow transformation
• Grow the transformed bacteria in the presence of substances such as ampicillin and X-gal

Question 3

Cloning vector

Answer 3

• An agent used to transfer DNA between cells
  
  • Plasmids are often used as vectors for they can be introduced into bacteria by transformation.

Question 4

amp^<em>R</em>gene

Answer 4

This gene gives a bacterium resistance against the antibiotic ampicillin.

Question 5

GFP gene

Answer 5

• This gene gives off a green fluorescence
  
  • it was originally attained from jellyfish

Question 6

lacZ gene

Answer 6

• This gene codes for an enzyme that normally breaks down lactose.
• This enzyme also breaks down the artificially made molecule, X-gal
• X-gal creates a blue product when broken down by lacZ enzyme.

Question 7

Restriction enzymes

Answer 7

• These recognize and cut DNA molecules that are foreign to the bacterium.
• They cut at specific restriction sites.

Question 8

Restriction fragment

Answer 8

A DNA segment that is the result of the cutting of DNA by an enzyme

Question 9

DNA ligase

Answer 9

• This is an enzyme that “catalyzes the formation of covalent bonds that close up the sugar-phosphate backbones of DNA strands” (Biology in Focus).
  
  • example - DNA ligase joins Okazaki fragments during replication

Question 10

Transformation

Answer 10

This occurs when bacteria absorb DNA from their surroudnings and incorporate this DNA into their own genome.

Question 11

Recombinant plasmids

Answer 11

Plasmids that contain the foreign DNA

Question 12

Who first discovered transformation?

How did this scientist come about this discovery?

Answer 12

Frederick Griffith

When trying to develop a vaccine against pneumonia, Griffith discovered transformation. He was experimenting with two strains of bacterium, one pathogenic, one nonpathogenic. He found that when he killed pathogenic bacteria and mixed their dead cells with living, nonpathogenic cells, some of those living cells became pathogenic