Because hybridization involves nucleotide bases and the separation and joining or reannealing of strands, several environmental factors can influence this process:

• Temperature: If the temperature is too high, the strands melt. If it is too low, they might be forced together.
• The pH: A pH that is too alkaline will cause the strands to separate; too acidic and they are forced together.
• The guanine to cytosine ratio (G:C ratio): Since this bond is stronger than the other nucleotide bonds, if the G:C ratio in the desired target strand is high, the separation process may take longer.