Recombinant DNA technology - amplification

Question 1

What is amplification of DNA?

Answer 1

Making more copies of a fragment of DNA.

Question 2

What two techniques are there of

amplifying DNA

Answer 2

n vitro amplification – (outside organisms) using the Polymerase Chain Reaction (PCR). In vivo amplification – (inside organisms) inserting the fragment into the genome of a host cell and culturing the host cell to produce more cells and therefore more copies of the gene.

Question 3

Describe the polymerase chain reaction.

Answer 3

Heat DNA to 90oC; Breaks hydrogen bonds and separates strands; Add primers; Add nucleotides; Cool; to allow binding of primers to nucleotides; increase temperature to the optimum for DNA polymerase; DNA polymerase joins adjacent DNA nucleotides together into a new strand; Repeat cycle many times;

Question 4

Why does the DNA replication in the PCR

eventually stop?

Answer 4

There are a limited number of primers and nucleotides available and these run out; so there is nothing to start/make the complimentary chains

Question 5

What is the role of primers in the PCR?

Answer 5

Short sequences of nucleotides with complementary bases to the end of the DNA fragments being copied by PCR. They bind to the start of the sequence to be copied and allow the DNA polymerase to join nucleotides to produce the complimentary strand.

Question 6

Why do primers only bind to specific

sequences?

Answer 6

Each primer have a specific base sequence; That is complementary to the start of the gene to be copied;

Question 7

What are the advantages of PCR over in vivo

amplification?

Answer 7

PCR can rapidly produce many copies. PCR does not require the use of living cells.