DNA manipulation.

Question 1

describe the process of PCR?

Answer 1

1. DENATURATION → the reaction mixture is heated to 90-90C, separating the double stranded DNA by breaking the H bonds b/w complementary bases.
2. ANNEALING → the reaction mixture is cooled to 55-60C allowing the primer to anneal (pair) with the DNA template. The cooler temp allows hydrogen bonds to form.
3. EXTENSION → reaction mixture is heated to 72C.This allows the Taq DNA polymerase (obtained from thermophilic bacteria found in hot springs) to synthesis the new strand of DNA → polymerase recognises the primer as the starting point for synthesis. It puts free floating nucleotides into the correct places along the DNA template to extend from the primer.

Question 2

give the advantages of PCR over in vivo techniques.

Answer 2

+ less labour intensive as u just set it to run
+ quicker –> takes a few cycles to produce lots of DNA samples over a few weeks.
+ can use low quality DNA

Question 3

give advantages of in vivo techniques over PCR.

Answer 3

• less prone to mutation by Taq Polymerase
• in vivo is less expensive.
• a longer piece of DNA can be clones in VIVI –> however PCR has a limited size for cloning.