2.4 DNA amplification using polymerase chain reaction.pptx

VCE Biology

1. 2.4 Polymerase Chain Reaction

2. 2.4 Polymerase Chain Reaction Learning Intentions Understand the amplification of DNA using polymerase chain reaction (PCR) Success Criteria explain the use of PCR. describe the process of DNA amplification.

3. Why PCR? So we isolate some DNA from a crime scene.. Studies on a single isolated piece of DNA are nearly impossible to conduct. So we need a way to amplify it: Polymerase Chain Reaction(PCR) PCR: a process to exponentially amplify a DNA sample

4. Steps in PCR

5. Step 1: denaturing The DNA sample needs to be separated into two separate strands. The separation happens by raising the temperature of the PCR mixture, causing the hydrogen bonds between the complementary DNA strands to break This occurs at approximately 94 °C for one minute.

6. Step 2: Primers anneal Short segments of single-stranded DNA, known as primers (forward and reverse sequences), are added. Primers bind to the target DNA sequences (at 3′ ends) and initiate DNA synthesis (polymerisation). These primers can bind to their complementary sequences on the single-stranded template DNA. This occurs at approximately 55 °C for two minutes.

7. Step 3: extending The polymerase enzyme (Taq polymerase) uses the primers as a starting point and extending the primers, synthesising the new strands of DNA by adding nucleotides. A supply of ‘free’ nucleotides must be available in order for the DNA polymerase to create a new strand of ‘complementary DNA’ to each of the single template strands. This occurs at 72 °C for one minute (the optimal temperature for Taq polymerase).

8. Step 4: REPEAT