DNA-DNA hybridisation
Encyclopedia
DNA-DNA hybridization generally refers to a molecular biology
technique that measures the degree of genetic similarity between pools of DNA
sequences. It is usually used to determine the genetic distance
between two species
. When several species are compared that way, the similarity values allow the species to be arranged in a phylogenetic tree
; it is therefore one possible approach to carrying out molecular systematics.
Charles Sibley
and Jon Ahlquist, pioneers of the technique, used DNA-DNA hybridization to examine the phylogenetic relationships of avians (the Sibley-Ahlquist taxonomy
) and primates. Critics argue that the technique is inaccurate for comparison of closely related species, as any attempt to measure differences between orthologous sequences between organisms is overwhelmed by the hybridization of paralogous sequences within an organism's genome. DNA sequencing and computational comparisons of sequences is now generally the method for determining genetic distance, although the technique is still used in microbiology to help identify bacteria.
The DNA of one organism is labeled, then mixed with the unlabeled DNA to be compared against. The mixture is incubated to allow DNA strands to dissociate and reanneal, forming hybrid double-stranded DNA. Hybridized sequences with a high degree of similarity will bind more firmly, and require more energy to separate them: i.e. they separate when heated at a higher temperature than dissimilar sequences, a process known as "DNA melting".
To assess the melting profile of the hybridized DNA, the double-stranded DNA is bound to a column and the mixture is heated in small steps. At each step, the column is washed; sequences that melt become single-stranded and wash off the column. The temperatures at which labeled DNA comes off the column reflects the amount of similarity between sequences (and the self-hybridization sample serves as a control). These results are combined to determine the degree of genetic similarity between organisms.
Molecular biology
Molecular biology is the branch of biology that deals with the molecular basis of biological activity. This field overlaps with other areas of biology and chemistry, particularly genetics and biochemistry...
technique that measures the degree of genetic similarity between pools of DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
sequences. It is usually used to determine the genetic distance
Genetic distance
Genetic distance refers to the genetic divergence between species or between populations within a species. It is measured by a variety of parameters. Smaller genetic distances indicate a close genetic relationship whereas large genetic distances indicate a more distant genetic relationship...
between two species
Species
In biology, a species is one of the basic units of biological classification and a taxonomic rank. A species is often defined as a group of organisms capable of interbreeding and producing fertile offspring. While in many cases this definition is adequate, more precise or differing measures are...
. When several species are compared that way, the similarity values allow the species to be arranged in a phylogenetic tree
Phylogenetic tree
A phylogenetic tree or evolutionary tree is a branching diagram or "tree" showing the inferred evolutionary relationships among various biological species or other entities based upon similarities and differences in their physical and/or genetic characteristics...
; it is therefore one possible approach to carrying out molecular systematics.
Charles Sibley
Charles Sibley
Charles Gald Sibley was an American ornithologist and molecular biologist. He had an immense influence on the scientific classification of birds, and the work that Sibley initiated has substantially altered our understanding of the evolutionary history of modern birds.Sibley's taxonomy has been a...
and Jon Ahlquist, pioneers of the technique, used DNA-DNA hybridization to examine the phylogenetic relationships of avians (the Sibley-Ahlquist taxonomy
Sibley-Ahlquist taxonomy
The Sibley-Ahlquist taxonomy is a bird taxonomy proposed by Charles Sibley and Jon Edward Ahlquist. It is based on DNA-DNA hybridization studies conducted in the late 1970s and throughout the 1980s....
) and primates. Critics argue that the technique is inaccurate for comparison of closely related species, as any attempt to measure differences between orthologous sequences between organisms is overwhelmed by the hybridization of paralogous sequences within an organism's genome. DNA sequencing and computational comparisons of sequences is now generally the method for determining genetic distance, although the technique is still used in microbiology to help identify bacteria.
The DNA of one organism is labeled, then mixed with the unlabeled DNA to be compared against. The mixture is incubated to allow DNA strands to dissociate and reanneal, forming hybrid double-stranded DNA. Hybridized sequences with a high degree of similarity will bind more firmly, and require more energy to separate them: i.e. they separate when heated at a higher temperature than dissimilar sequences, a process known as "DNA melting".
To assess the melting profile of the hybridized DNA, the double-stranded DNA is bound to a column and the mixture is heated in small steps. At each step, the column is washed; sequences that melt become single-stranded and wash off the column. The temperatures at which labeled DNA comes off the column reflects the amount of similarity between sequences (and the self-hybridization sample serves as a control). These results are combined to determine the degree of genetic similarity between organisms.