Unit 7- Manipulating DNA

Question 1

Where do restriction endonucleases cut DNA?

Answer 1

Through the phosphodiester bonds

Question 2

How are blunt end and sticky ends formed?

Answer 2

Blunt ends- when the restriction endonucleases cuts straight across both chains
Sticky ends- when enzymes make a staggered cut

Question 3

What are sticky ends?

Answer 3

When restriction endonucleases make a staggered cut through DNA, revealing short stretches of single stranded DNA that will anneal to a complementary base pair sequence

Question 4

What is a recognition sequence?

Answer 4

The short section of DNA that is complementary to the restriction endonucleases active site so is cut

Question 5

What pattern do recognition sequences display?

Answer 5

Palindromic

Eg (GAATTC, which reversed is CTTAAG)

Question 6

What is the purpose of DNA ligase.

Answer 6

Reform the phosphodiester bond in the backbone of DNA

Question 7

Where are reverse transcriptase enzymes found naturally

Answer 7

In retroviruses

Question 8

What does reverse transcriptase do?

Answer 8

It synthesises DNA which codes for exactly 1 protein from mRNA

Question 9

Describe the process of how an artificial gene is made from mRNA

Answer 9

• mRNA (no introns) is extracted from cells and mixed with reverse transcriptase and nucleotides
• the enzyme copies the mRNA to make double stranded copied DNA (cDNA)
• cDNA is 1 gene long however much shorter than normal DNA as there are no introns to assist in structure etc
• cDNA is an artificial gene

Question 10

What are the benefits of using reverse transcriptase?

Answer 10

• makes genes without introns (the exact sequence)
• copy of the gene is still stable
• as its only a short strand of DNA it’s easier to insert into a plasmid