1.2) Replication of DNA

Question 1

• What is the enzyme used to replicate DNA prior to cell division and what does it need?

Answer 1

➜ DNA Polymerase 🧬

• DNA Polymerase
  ➞ needs primers to start replication.
• A ‘primer’
  ➞ is a short strand of nucleotides which binds at the 3’ end of the template DNA strand 3️⃣
  ↳ allowing DNA polymerase to add nucleotides ➕.

Question 2

• What are the requirements for DNA replication?

Answer 2

1. ➜ Parental DNA
2. ➜ Enzymes (DNA Polymerase + Ligase)
3. ➜ Free DNA nucleotides
4. ➜ ATP
5. ➜ Primers

Question 3

• What is the 1st step of DNA replication?

Answer 3

UNWINDS:
1. DNA
➞ unwinds and hydrogen bonds between bases are broken
↳ to form 2 template strands.

UNZIPS:
2. The 2 strands of DNA
➞ separate (unzip) and the bases are exposed.`

Question 3

• What is the 2nd step of DNA replication?

Answer 3

PRIMER & DNA POLYMERASE:
1. a PRIMER
➞ (a short strand of complementary bases)
↳ acts as a “starting point” for DNA Polymerase.

2. DNA POLYMERASE
   ➞ (enzyme)
   ↳ adds complementary DNA nucleotides to the deoxyribose 3’ end of the new strand that forms.

LEADING & LAGGING STRANDS:
3. DNA polymerase
➞ can only add nucleotides in one direction ➡️
↳ resulting in one strand (leading strand) being replicated continuously.

4. The other strand (lagging strand)
   ➞ is replicated in fragments.
   ↳ These fragments are then joined together by another enzyme (LIGASE)

Question 4

• What is the 3rd step of DNA replication?

Answer 4

BONDS:
1. Weak hydrogen bonds
➞ are formed between the complementary base pairs.

2. A strong bond
   ➞ is formed between the deoxyribose sugar and phosphates of neighbouring nucleotides.
   ↳ = strong sugar-phosphate backbone.

Question 5

• What is the overview of the LEADING STRAND?

Answer 5

• 1.) ➞ DNA unzips 🧬
• 2.) ➞ PRIMER attaches to the 3’ END of the parental DNA 3️⃣
• 3.) ➞ Free nucleotides align with the complementary base pairs 🔬
• 4.) ➞ DNA POLYMERASE binds the free nucleotides to the DNA primer’s 3’end 3️⃣
• 5.) ➞ DNA polymerase works continuously to form the leading strand 🧬

Question 6

• What is the overview of the LAGGING STRAND?

Answer 6

• 1.) ➞ DNA unzips 🧬
• 2.) ➞ Primer attaches at the replication fork 🍴
• 3.) ➞ Free nucleotides align with complementary base pairs 🔬
• 4.) ➞ DNA POLYMERASE binds the free nucleotides to the PRIMER’S 3’ END 3️⃣
• 5.) ➞ This process is repeated further up the parental DNA strand forming fragments 🧬
• 6.) ➞ The fragments are joined together by DNA ligase⚡

Question 7

• PAST PAPER (REVIEW) QUESTIONS:

1. Which of the following statements about DNA replication is correct?
   a. ➜Polymerase adds nucleotides to the 3’ end of a DNA strand
   b. ➜Polymerase adds nucleotides to the 5’ end of a DNA strand
   c. ➜Ligase adds nucleotides to the 3’ end of a DNA strand
   d. ➜Ligase adds nucleotides to the 5’ end of a DNA strand
2. Explain why DNA replication must take place before a cell divides.

Answer 7

1. ➜ A
2. ➜ It must take place before a cell divides in order to maintain the diploid chromosome complement (full chromosome complement)

Question 8

• What is PCR?

Answer 8

• PCR (‘polymerase chain reaction’)
• to amplify DNA ‘in vitro’ 🔍🧪
• using complementary primers for specific target sequences 🧬

Question 9

• What is step 1 in the PCR process?

Answer 9

1. HEATING: 🔥
   * Target DNA, DNA polymerase, primers and nucleotides are mixed together.
   * The mixture is then heated to between 92°C and 98°C
   to separate the DNA strands 🔥
   * (H-bonds are broken between bases to separate the two strands)

Question 10

• What is step 2 in the PCR process?

Answer 10

2. ANNEALING: ✨
   * Temperature is reduced to between 50°C and 56°C
   * To allow primers to anneal (forming H-bonds) to their complementary sequence on the separated strands 🧬
   * The primers (about 20) will bind to both strands of the target sequence of DNA

Question 11

• What is step 3 in the PCR process?

Answer 11

3. AMPLIFICATION: 🔍
   * Temperature is raised to between 70°C and 80°C
   * To allow the heat tolerant Taq polymerase to add nucleotides to the 3’ ends of the primer 🔥
   * It extends them into new complementary strands
   (replicating the region of DNA)

Question 12

• What can PCR-amplified DNA do?

Answer 12

1. help solve crimes 🚨
2. settle paternity suits ⚖️
3. diagnose genetic disorders 🧬

Question 13

• PAST PAPER (REVIEW) QUESTIONS:
  1. State the function of PCR.

2. Describe what happens to the DNA in Stage 1.
3. Short sections of DNA called primers are involved in Stage 2.
   ➞ State what happens to these primers during Stage 2.
4. Suggest why the temperature is increased during Stage 3.

Answer 13

1. PCR is used to amplify DNA ‘in vitro’ using complementary primers for specific target sequences.
2. The target DNA, DNA polymerase, primers and nucleotides are mixed together which is then heated to between 92 and 98°C which separates the DNA strands (H-bonds are broken).
3. The primers (about 20) will anneal to both strands of the target sequence of DNA.
4. The increased temperature allows the Taq polymerase to add nucleotides to the 3’ ends of the primer.