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Describe and explain how the polymerase chain reaction (PCR) is used to amplify a DNA fragment - AQA - A-Level Biology - Question 8 - 2021 - Paper 2
Question 8
Describe and explain how the polymerase chain reaction (PCR) is used to amplify a DNA fragment. Figure 9 shows the number of DNA molecules produced using a PCR. ... show full transcript
Worked Solution & Example Answer:Describe and explain how the polymerase chain reaction (PCR) is used to amplify a DNA fragment - AQA - A-Level Biology - Question 8 - 2021 - Paper 2
Step 1
Describe and explain how the polymerase chain reaction (PCR) is used to amplify a DNA fragment.
Answer
The polymerase chain reaction (PCR) is a widely used technique in molecular biology to amplify specific DNA fragments. The process involves several key steps:
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Preparation of Components: The PCR requires a DNA template (the fragment to be amplified), DNA polymerase (commonly Taq polymerase), primers (short DNA sequences complementary to the target DNA), and nucleotides.
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Denaturation: The reaction mixture is heated to around 95°C, causing the double-stranded DNA to denature. This step breaks the hydrogen bonds between the complementary bases, resulting in two separate strands.
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Annealing: The temperature is then lowered to approximately 50-65°C, allowing the primers to anneal (bind) to the specific sequences on the single-stranded DNA templates. This step is crucial for ensuring that the amplification is specific to the target region.
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Extension: Finally, the temperature is raised to around 72°C, the optimal temperature for the DNA polymerase. During this step, the polymerase synthesizes new DNA strands by adding nucleotides to the primers, thus extending the DNA strands. This cycle of denaturation, annealing, and extension is repeated for multiple cycles (usually 25-35), leading to exponential amplification of the target DNA fragment.
Step 2
Explain the shape of the curve in Figure 9.
Answer
The shape of the curve in Figure 9 can be explained in two main phases:
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Initial Phase: At the beginning of the PCR cycles, the number of DNA molecules increases slowly. This is due to the limited amount of primers and nucleotides, resulting in a low number of doublings per cycle.
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Exponential Phase: After several cycles, the number of DNA molecules begins to double with each cycle, producing an exponential increase in the quantity of DNA.
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Plateau Phase: Eventually, the curve levels off and reaches a plateau. This plateau occurs because the PCR components (such as primers and nucleotides) become depleted, and the reaction system reaches a saturation point. At this stage, the enzymatic activity may also decrease due to enzyme denaturation.