DNA Hybridization

Evolution deals with heritable changes in populations over time. Because DNA is the molecule of heredity, evolutionary changes will be reflected in changes in the base pairs in DNA. Two species that have evolved from a common ancestor will have DNA that has very similar base pair sequences. The degree of relatedness of two species can be estimated by examining how similar their base pair sequences are. One method of assessing relatedness uses hybridization of DNA.

In the molecular genetic technique of hybridization of DNA, single strands of DNA from two different species are allowed to join together to form hybrid double helices. These hybrid segments of DNA can be used to determine the evolutionary relatedness of organisms by examining how similar or dissimilar the DNA base pair sequences are.

The technique of DNA hybridization is based on two principles: the first, that double strands of DNA are held together by hydrogen bonds between complementary base pairs, and the second is that the more closely related two species are, the greater will be the number of complementary base pairs in the hybrid DNA. In other words, the degree of hybridization is proportional to the degree of similarity between the molecules of DNA from the two species.

Hybridization of DNA is accomplished by heating strands of DNA from two different species to 86° C [186.8° F]. This breaks the hydrogen bonds between all complementary base pairs. The result is many single-stranded segments of DNA. The single-stranded DNA from both species is mixed together and allowed to slowly cool. Similar strands of DNA from both species will begin to chemically join together or re-anneal at complementary base pairs by reforming hydrogen bonds.

The resulting hybrid DNA is then reheated and the temperature at which the DNA once again becomes single-stranded is noted. Because one cannot observe DNA separating, another technique must be used simultaneously with heating to show when separation has occurred. This technique employs the absorption of UV light by DNA. Single strands of DNA absorb UV light more effectively than do double strands. Therefore, the separation of the DNA strands is measured by UV light absorption; as more single strands are liberated, more UV light is absorbed.

The temperature at which hybrid DNA separation occurs is related to the number of hydrogen bonds formed between complementary base pairs. Therefore, if the two species are closely related, most base pairs will be complementary and the temperature of separation will be very close to 86° C [186.8° F]. If the two species are not closely related, they will not share many common DNA sequences and fewer complementary base pairs will form. The temperature of separation will be less than 86° C [186.8° F] because less energy is required to break fewer hydrogen bonds. Using this type of information, a tree of evolutionary relationships based on the separation temperature of the hybrid helices can be generated.

See also Evolution and evolutionary mechanisms; Evolutionary origin of bacteria and viruses

DNA-DNA hybridization A technique that is used to compare DNA from two different species, to locate or identify nucleotide sequences, and to establish the effective in vitro transfer of nuclear material to a new host. A single strand of DNA from one source is bound to a special filter to which is added a single strand of radioactively labelled DNA from a different source. Complementary base pairing between homologous sections of the two DNAs results in double-stranded hybrid sections that remain bound to the filter, whereas single-strand sections are washed away. The amount of radioactivity remaining on the filter, compared with the amount washed away, gives a measure of the number of nucleotide sequences that the radioactive DNA and the original DNA share in common.

DNA hybridization A method of determining the similarity of DNA from different sources. Single strands of DNA from two sources, e.g. different bacterial species, are put together and the extent to which double hybrid strands are formed is estimated. The greater the tendency to form these hybrid molecules, the greater the extent of complementary base sequences, i.e. gene similarity. The method is one way of determining the genetic relationships of species.