Gene Technology

Question 1

What are the limitations of PCR?

Answer 1

It is hard to amplify fragments larger than 15kb. The flaking sequence must be known in order to attach the primer

Question 2

Where re restriction endonuclease harvested from?

Answer 2

The restriction endonuclease come from bacteria

Question 3

What stops the bacterial enzymes from breaking down their own DNA?

Answer 3

The enzyme ECO R1 stops endonuclease from cutting its own DNA

Question 4

What is used as a vector in PCR?

Answer 4

Plasmids are used a a vector

Question 5

What are the main steps of PCR?

Answer 5

DNA fragment is extracted by heating it to 94 degrees and cutting using a restriction endonuclease.

Question 6

How are E.coli cells prepared?

Answer 6

The E.coli is added to calcium chloride at 50mM, the it is heat shocked at 42 degrees. The plasmids enters the E.coli

Question 7

How can replicating plating be used to identify the recombinant DNA and the self-replicating?

Answer 7

The gene is inserted into the tetracyclin gene, which breaks it. All vectors are added to amplicilin plate. Them tested in tetracylin, ones that do not survive are the recombinant DNA