DNA Replication and PCR

Question 1

What is a replication fork and what is the role of Helicase in its formation?

Answer 1

The enzyme Helicase begins replication by unwinding the double helix and separating the strands by breaking the weak hydrogen bonds between complementary bases on each strand. This creates an area called the replication fork.

Question 2

How many carbons are present in the sugar contained within a DNA nucleotide?

Answer 2

5.

Question 3

What is a Nucleosome?

Answer 3

The strand of DNA wrapped around histones.

Question 4

In which direction does DNA replication occur and why?

Answer 4

5’ - 3’ as DNA Polymerase is only able to work in that direction.

Question 5

What does PCR stand for and can you explain its significance?

Answer 5

PCR stands for Polymerase Chain Reaction and it is used to amplify DNA, allows testing on certain sequences of DNA for a multitude of purposes.

Question 6

Explain the difference between the Leading Strand and the Lagging Strand.

Answer 6

The leading strand can be replicated continuously since it runs in the same direction as DNA replication occurs. The lagging strand is replicated in fragments, known as Okazaki fragments, since it runs in the opposite direction due to a double helix being antiparallel. This is known as discontinuous replication.

Question 7

What is meant by the term “semi-conservative” in the context of DNA replication?

Answer 7

The new Double Helix keeps half of the original DNA

Question 8

What is the role of DNA polymerase and DNA ligase in DNA replication?

Answer 8

DNA polymerase is used to bind to free DNA nucleotides to DNA template. DNA ligase is used to fuse Okazaki fragments together.

Question 9

Name the chemical bonds that form between complementary DNA base pairs

Answer 9

Hydrogen Bonds

Question 10

How many bonds bind the DNA base pairs together? (A-T and C-G)

Answer 10

Two Chemical bonds bind A-T and three chemical bonds bind C-G.

Question 11

What are the bonds that form between the Phosphate group and Deoxyribose sugar?

Answer 11

Phosphodiester bonds

Question 12

What is an Okazaki fragment?

Answer 12

An Okazaki fragment is the fragments that make up the lagging strand of DNA.

Question 13

What enzyme is used for the replication of DNA during PCR?

Answer 13

DNA (taq) polymerase

Question 14

At the commencement of PCR there are 4 molecules of template DNA. How many molecules of DNA would be present after 5 cycles of PCR?

Answer 14

4x2^5=128

Question 15

Why does the enzyme used to amplify DNA in PCR not denature in high heats?

Answer 15

It has an optimum temperature of 72oC and therefore doesn’t denature in high heats such as 95oC.

Question 16

Name the 3 stages of PCR and what happens at each.

Answer 16

Denaturing, Annealing, and Elongation. In the denaturing stage the DNA is heated to 92oC – 98oC for 15-30 seconds to denature DNA strands – separating the strands.  In the Annealing stage Cool (50oC-65oC) to bind primers (anneal) to template. In the Elongation stage the value of PCR presents itself by increasing the number of DNA fragments.

Question 17

What is required for DNA replication?

Answer 17

DNA template, Primer, Supply of Nucleotides, Enzyme, DNA Polymerase, Ligase and ATP.

Question 18

What are 3 practical uses of PCR?

Answer 18

Solve crime/forensics, Settle paternity suits, Diagnose genetic disorders.

Question 19

What is Gel Electrophoresis?

Answer 19

Used to separate different DNA fragments on the basis of size. Samples of DNA (typically with a dye) are loaded into wells at one end and an electric current drags them through the gel. This works because DNA fragments are negatively charged and therefore move towards the positive electrode. Smaller fragments move faster than large ones.