International A Level Biology / Decomposition and forensics
The polymerase chain reaction (PCR)
Edexcel International A Level Biology (YBI11), chapter 19: Decomposition and forensics, specification point 6.17. Free lesson.
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PCR amplifies DNA by repeatedly separating its strands and using them as templates to make new strands. It is useful when a sample contains too little DNA to analyse, such as a single hair from a crime scene.
What goes into a PCR tube
PCR uses one small tube placed in a thermal cycler, a machine that heats and cools it repeatedly. Each ingredient has a different job. DNA sample: the DNA to copy. Each strand acts as a template, or pattern, for building a new strand. Primers: short single strands of DNA that bind to the start of the section being copied. One binds to each DNA strand. Free nucleotides: DNA building blocks containing the bases A, T, C and G. They are the raw material for new strands. Taq polymerase: a DNA polymerase from the hot spring bacterium *Thermus aquaticus*. It joins nucleotides…
How one PCR cycle makes two DNA molecules
A cycle has three temperature stages. The exact temperatures can vary slightly, so use the temperatures given in a question. After one cycle, there are two DNA molecules, each made from one old strand and one new strand. At 95 °C, heat breaks the hydrogen bonds between the bases. The two DNA strands separate. This is similar to the strand-separating role of helicase in a cell. At about 55 °C, the primers anneal, which means they bind to the strands by complementary base pairing. The strands have different base sequences, so each needs its own primer. At 72 °C, the optimum…
Taq's role and why it is used
Be clear about the difference between what Taq does and why it is chosen. Its role is to build new DNA strands. It is chosen because it can keep working through the repeated high-temperature stages. Taq joins free nucleotides with phosphodiester bonds, making a strand complementary to the template. Taq is heat stable, so it is not denatured at 95 °C. An ordinary enzyme would be denatured in the first cycle.
How to calculate the number of DNA copies
If every DNA molecule is copied in each cycle, the number doubles each time. This makes the number of copies increase very quickly. Use molecules = N × 2ⁿ. N is the number of DNA molecules at the start. n is the number of cycles. If you are given the total time and the time for one cycle, first calculate the number of cycles. Then calculate 2ⁿ and multiply by N.