DNA Hybridisation

DNA hybridisation is the binding of a single strand of DNA (or RNA) to another single strand with a complementary base sequence. It is how a DNA probe finds its target.1. The DNA sample to be tested is heated to around 95 °C. This breaks the hydrogen bonds between complementary bases, so the double helix separates into single strands.
2. The labelled probe is added.
3. The mixture is cooled. Complementary bases pair up and new hydrogen bonds form. If the sample contains a sequence complementary to the probe, the probe binds to it, forming a hybrid double strand of probe and sample DNA.
4. The DNA is washed to remove any probe that has not bound.
5. The label is detected, by fluorescence under UV light or by autoradiography. If the label is present, the target sequence is present.Hybridisation depends on how closely the sequences match. The more complementary base pairs there are, the more hydrogen bonds form and the more stable the hybrid is. Under strict conditions, such as a higher temperature during washing, only a perfectly complementary probe stays bound, so even a single base difference between alleles can be detected.Hybridisation can also show how closely related two species are. DNA from both species is mixed, separated into single strands and allowed to hybridise, then the temperature needed to separate the hybrid strands is measured. The more similar the base sequences, the more hydrogen bonds there are, so a higher temperature is needed.