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An oligonucleotide is a short chain of DNA or RNA molecules that has many uses in molecular biology and medicine. It is used as a probe to screen for diseases, viral infections, and to identify genes in molecular biology experiments. It is also used as a primer in a type of DNA sequencing.
To understand an oligonucleotide, it helps to understand the structure of DNA. DNA molecules are very long coils of two strands, made of four different nucleotide base units, arranged in varying orders. Each unit has a complementary base it will bind, so each strand has an opposite set of bases binding it. These bases can form a great variety of different combinations, and it is the combination of bases that gives the genetic code. DNA is transcribed to produce messenger RNA (mRNA), which is then translated to produce proteins.
Oligonucleotides are identified by their chain length. For instance, an oligonucleotide that is ten nucleotide bases long would be called a ten mer. They are generally synthesized chemically, and the type of synthesis limits the chain length to less than 60 bases long.
In a type of DNA sequencing known as dideoxy sequencing, oligonucleotides are used as a primer, so that the enzyme that makes the DNA will have a template to work from. Single-stranded DNA is used, and an oligonucleotide that is complementary to the DNA strand is synthesized using an automated machine. The DNA polymerase that synthesizes the DNA continues adding onto the primer, and synthesizes the opposite strand of DNA from it. This reaction produces double-stranded DNA.
A more recent use of oligonucleotides as primers is in the polymerase chain reaction (PCR) that is used to amplify small fragments of DNA. This technique has highly practical uses, such as in forensics and paternity testing. It has also revolutionized research in medicine and the biological sciences, since it is often used in genetic engineering experiments.
A series of oligonucleotide probes is frequently used to isolate genes from a library of genes or complementary DNAs (cDNAs). cDNA libraries consist of double-stranded DNA, in which one strand is derived from an mRNA strand and the other is complementary to it. Such libraries have the advantage that they do not have the gaps that are frequently found in genes from higher organisms.
The structure of genes from many organisms is known, due to sequencing projects, and is publicly available. If one wants to clone a gene from another organism, one can see what is known about the gene in other organisms, and design probes based on common areas in those sequences. Researchers then have a series of oligonucleotide probes synthesized that take into account possible variations at the common area. They screen the library with those probes and look for oligonucleotides that bind. Many genes have been identified in this manner.
An antisense oligonucleotide contains a single strand of RNA or DNA that is the complement of a sequence of interest. Once a particular gene coding for a protein has been cloned, antisense RNA is frequently used to block its expression by binding to the mRNA that would synthesize it. This lets researchers determine the effects on the organism, when it does not make that protein. Antisense oligonucleotides are also being developed as novel types of drugs to block toxic RNAs.
Microarray chips have been another area in which oligonucleotides have been of great utility. These are glass slides, or some other matrix, that have spots containing thousands of different DNA probes — in this case made of oligonucleotides. They are a very efficient way to test for changes in a number of different genes at once. The DNA is bound to a compound that changes color, or fluoresces, if complementary DNA binds to it, so the spots change color if there is a reaction with the test DNA.
Some of the things oligonucleotide microarrays are used for include screening for genetic diseases. For example, there are small probes that represent the activity of genes involved in breast cancer, BRCA1 and BRCA2. It is possible to find out whether a woman has a mutation in one of these genes, and analyze it further to see if she has a predisposition for breast cancer.
There is a microarray called the ViroChip that has probes for about 20,000 genes from various pathogenic viruses that have been sequenced. Body secretions, such as phlegm, can be analyzed with the chip, which can often identify what type of virus a person is infected with. Identifying viral infections can be quite difficult, since the symptoms are often similar with different types of viruses.
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