A3.1 DNA Barcoding - student reading
There have been many sightings of mythical beasts including Bigfoot, Yeti, Sasquatch and Loch Ness Monster.
Little evidence exists to illustrate their existence other than folklore.
Sometimes, physical evidence is left behind like hair or faeces. With DNA barcodes we can help to identify these unknown creatures. Below features a table from research conducted by Sykes et al. displaying DNA barcode results on evidence from supposed sightings of Bigfoot/Yeti.
Table source - http://rspb.royalsocietypublishing.org/content/281/1789/20140161 CC-BY
The Need for Barcoding
These days, identification of organisms is becoming increasingly important as a measurement of biodiversity in the face of habitat destruction and global climate change. There is no consensus on how many species exist on this planet, but the estimation of extinction rates is about 1 species per 100-1000 million species. The more information we have about the presence of absence of species within a habitat the better we will be able to avoid extinctions.
Classification from Linné’s to the twentieth century was mostly performed by morphological differences.Molecular biology and DNA technologies have revolutionised the classification system of living things especially in providing the ability to match relatedness of these species. DNA barcoding, like the name implies, seeks to use DNA markers to identify organisms.
The origin of DNA barcodes
Watch this short video interview with about the origin and uses of DNA barcodes. Video from the DNA Learning Center
DNA barcoding uses DNA markers to identify organisms. But what DNA should be used?
Since the goal of barcoding is to define specific organisms, discrimination is the primary objective.
There has to be a difference in the DNA sequence between species.
However, when there is some universality in the DNA used for discrimination the same locus could identify a very wide range of genomes. There must be some universality in the DNA marker.
While discrimination is about the uniqueness of sequences, universality seeks to use a single methodology, using one set of PCR primers for the same distinct region. This makes the technique more widely useful, and easier to use.
If some region of DNA has absolutely no sequence deviation between species, this has great universality but poor discrimination. But if a sequence has very low sequence similarity, this is great for discrimination but has absolutely no universality and can not be amplified with the same set of primers.
This is clearly illustrated by the three examples.
They each show DNA sequences from five different species. Each row represents a species:
- A case where there is universality for designing primers, but not an area where discrimination can occur.
- A case where discrimination of different organisms can occur, but the lack of similarity in sequence would make it difficult to design primers. That is, the lack of universality in sequence would prevent its use in many species.
- Enough variability in these sequences gives us the ability to discriminate between species.
The high similarity provides the universality required to design DNA primers that will work in many species.
Different DNA barcodes are used for animals, plants and prokaryotes
Sometimes, species are so similar for one DNA barcode sequence that a second marker is required. Scientists sometimes use a third, or fourth set of DNA markers to aid in increased discrimination.
In animals, the most commonly used barcode is the mitochondrial gene, Cytochrome Oxidase I. Since all animals have mitochondria and have this mitochondrial gene, it offers high universality. It is a DNA locus that is easy to amplify with one single technique, and each species has enough sequence deviation to discriminate between them.
Animal mitochondrial genomes vary from 16kb-22kb in size. However, plants, fungi, and protists have wildly different and larger mitochondrial genomes. For plants, we use a chloroplast gene, ribulose-bisphosphate carboxylase large subunit (rbcL)
Prokaryotes are often discriminated by their 16s rRNA gene (that's the gene for 16s ribosomes) while eukaryotes can be identified by 18s rRNA (They have 18s ribosomes.)
Hybrid organisms
A single DNA barcode will not create a clear picture of species identity in the case of hybrid animals (mules, ligers, coydogs, etc.). Sometimes, closely related species as well as hybrids are also indistinguishable by a single barcode. By developing multiple barcodes there is flexibility in the system, if one barcode doesn't work, there may be another that does.
Measuring Biodiversity using DNA
The technologies of molecular genetics, data processing, and data storage are maturing to the point where cataloguing the planet’s species in an accessible way is close to feasible.
DNA barcoding is a taxonomic method that uses a short genetic marker in an organism’s DNA to identify it as belonging to a particular species. The most commonly-used barcode region for animals, at least, is a segment of approximately 600 base pairs of the mitochondrial gene cytochrome oxidase I (COI).
Applications of DNA barcoding include,
- identifying plant leaves (even when flowers or fruit are not available),
- identifying insect larvae (which may have fewer diagnostic characters than adults and are frequently less well-known),
- identifying the diet of an animal (based on its stomach contents or faeces), and
- identifying products in commerce (for example, herbal supplements or wood).
Rapid, mass-sequencing machines make the molecular genetics portion of the work relatively inexpensive and quick. Computer resources store and make available the large volumes of data. Projects are currently underway to use DNA barcoding to catalogue museum specimens, which have already been named and studied, as well as testing the method on less studied groups. There are significant numbers of undescribed species that looked too much like sibling species to previously be recognized as different. These now can be identified with DNA barcoding.
Numerous computer databases now provide information about named species and a framework for adding new species. However, as already noted, at the present rate of description of new species, it will take close to 500 years before the complete catalogue of life is known. Many, perhaps most, species on the planet do not have that much time.
There is also the problem of understanding which species known to science are threatened and to what degree they are threatened. This task is carried out by the non-profit IUCN (International Union for Conservation of Nature) which maintains the Red List: an online listing of endangered species categorized by taxonomy, type of threat, and other criteria. The Red List is supported by scientific research. In 2011, the list contained 61,000 species, all with supporting documentation.
- DNA barcoding is a method used in taxonomy.
- Identification of species can be done from environmental DNA in a habitat
- A short genetic marker in an organism’s DNA is used to identify it as belonging to a particular species.
- Barcoding allows us to classify organisms that would otherwise be difficult to identify, such as in situations where only part of an organism is available, or is too immature to identify by studying its morphology.
- This method allows the biodiversity of habitats to be investigated rapidly.
- At the present rate of description of new species, it will take close to 500 years before the complete catalogue.
- Even with barcoding, it is difficult to know which species are threatened with extinction, a task carried out by the non-profit IUCN (International Union for Conservation of Nature).