Polymerase Chain Reaction (PCR): Amplifying DNA
Definition
PCR (Polymerase Chain Reaction)
A technique that amplifies DNA, creating millions of copies of a specific DNA sequence in just a few hours.
This allows scientists to study tiny DNA samples from crime scenes, fossils, or other sources.
Key Components of PCR
- Template DNA: The DNA you want to copy.
- Primers: Short DNA sequences that bind to the template, marking the starting point for replication.
- Taq Polymerase: A heat-resistant enzyme that synthesizes new DNA strands.
- Nucleotides (dNTPs): The building blocks of DNA.
- Thermal Cycler: A machine that precisely controls temperature changes.
- Primers are essential because Taq polymerase can’t start building DNA without them.
- They are designed to match the beginning and end of the target sequence.
Steps of PCR (DNA Amplification)
PCR consists of a cycle of three main steps, repeated 20–40 times to exponentially amplify the target DNA sequence.
- Sample Preparation
- The DNA sample is extracted and purified.
- Template DNA (the DNA to be copied) is prepared along with other PCR components.
- Denaturation (~94-96°C) → Fragmentation
- The DNA is heated to ~95°C.
- This breaks the hydrogen bonds between complementary base pairs, separating the double-stranded DNA into two single strands.
- These single strands serve as templates for new DNA synthesis.
- Annealing (~50-65°C) → Primer Annealing
- The temperature is lowered to 50–65°C.
- Short DNA sequences called primers bind (anneal) to complementary sequences on each single-stranded DNA template.
- Primers mark the starting point for DNA synthesis and are essential because Taq polymerase cannot start building DNA without them.
- Extension (~72°C) → DNA Synthesis
