Because primers are designed to have low complementarity to each other, they may anneal (step I in the figure) only at low temperature, e.g. room temperature, such as during the preparation of the reaction mixture. Although DNA polymerases used in PCR are most active around 70°C, they have some polymerizing activity also at lower temperatures, which can cause DNA synthesis from primers after annealing to each other. Several methods have been developed to prevent PDs formation until the reaction reaches working temperature (60-70°C), and these include initial inhibition of the DNA polymerase, or physical separation of reaction components reaction until the reaction mixture reaches the higher temperatures. These methods are referred to as hot-start PCR.
Wax: in this method the enzyme is spatially separated from the reaction mixture by wax that melts when the reaction reaches high temperature.
Slow release of magnesium: DNA polymerase requires magnesium ions for activity, so the magnesium is chemically separated from the reaction by binding to a chemical compound, and is released into the solution only at high temperature.