Open main menu

Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications in genetic testing, research, and forensics. Commonly made in the laboratory by solid-phase chemical synthesis, these small bits of nucleic acids can be manufactured as single-stranded molecules with any user-specified sequence, and so are vital for artificial gene synthesis, polymerase chain reaction (PCR), DNA sequencing, library construction and as molecular probes. In nature, oligonucleotides are usually found as small RNA molecules that function in the regulation of gene expression (e.g. microRNA), or are degradation intermediates derived from the breakdown of larger nucleic acid molecules.

Oligonucleotides are characterized by the sequence of nucleotide residues that make up the entire molecule. The length of the oligonucleotide is usually denoted by "-mer" (from Greek meros, "part"). For example, an oligonucleotide of six nucleotides (nt) is a hexamer, while one of 25 nt would usually be called a "25-mer". Oligonucleotides readily bind, in a sequence-specific manner, to their respective complementary oligonucleotides, DNA, or RNA to form duplexes or, less often, hybrids of a higher order. This basic property serves as a foundation for the use of oligonucleotides as probes for detecting DNA or RNA. Examples of procedures that use oligonucleotides include DNA microarrays, Southern blots, ASO analysis, fluorescent in situ hybridization (FISH), PCR, and the synthesis of artificial genes. Oligonucleotides are also indispensable elements in antisense therapy.[1][2]

Oligonucleotides composed of 2'-deoxyribonucleotides (oligodeoxyribonucleotides) are fragments of DNA and are often used in the polymerase chain reaction, a procedure that can greatly amplify almost any small amount of DNA. There, the oligonucleotide is referred to as a primer, allowing DNA polymerase to extend the oligonucleotide and replicate the complementary strand.


Oligonucleotides are chemically synthesized using building blocks, protected phosphoramidites of natural or chemically modified nucleosides or, to a lesser extent, of non-nucleosidic compounds. The oligonucleotide chain assembly proceeds in the direction from 3'- to 5'-terminus by following a routine procedure referred to as a "synthetic cycle". Completion of a single synthetic cycle results in the addition of one nucleotide residue to the growing chain. A less than 100% yield of each synthetic step and the occurrence of side reactions set practical limits of the efficiency of the process so that the maximum length of synthetic oligonucleotides hardly exceeds 200 nucleotide residues. HPLC and other methods can be used to isolate products with the desired sequence.

Antisense oligonucleotidesEdit

Antisense oligonucleotides are single strands of DNA or RNA that are complementary to a chosen sequence. In the case of antisense RNA they prevent protein translation of certain messenger RNA strands by binding to them. Antisense DNA can be used to target a specific, complementary (coding or non-coding) RNA. If binding takes place this DNA/RNA hybrid can be degraded by the enzyme RNase H. The use of morpholino-antisense oligonucleotides for gene knockdowns in vertebrates, which is now a standard technique in developmental biology and is used to study altered gene expression and gene function, was first developed by Janet Heasman using Xenopus. .[3] The antisense oligonucleotides have also been used to inhibit influenza virus replication in cell lines.[4][5]

DNA microarrayEdit

One subtype of DNA microarrays can be described as substrates (nylon, glass, etc.) to which oligonucleotides have been bound at high density. There are a number of applications of DNA microarrays within the life sciences.

Chemical analysisEdit


Alkylamides can be used as chromatographic stationary phases.[6] Those phases have been investigated for the separation of oligonucleotides.[7]

Mass spectrometry

A mixture of 5-methoxysalicylic acid and spermine can be used as a matrix for oligonucleotides analysis in MALDI mass spectrometry.[8]

See alsoEdit

  • Aptamers, oligonucleotides with important biological applications
  • Morpholinos, oligos with non-natural backbones, which do not activate RNase-H but can reduce gene expression or modify RNA splicing
  • Polymorphism, the appearance in a population of the same gene in multiple forms because of mutations; can often be tested with ASO probes
  • CpG Oligodeoxynucleotide, an ODN with immunostimulatory properties
  • Polypurine reverse-Hoogsteen hairpins, PPRHs, oligonucleotides that can bind either DNA or RNA and decrease gene expression.


  1. ^ Weiss, B., ed. (1997). Antisense Oligodeoxynucleotides and Antisense RNA : Novel Pharmacological and Therapeutic Agents. Boca Raton, Florida: CRC Press
  2. ^ Weiss, B., Davidkova, G., and Zhou, LW (March 1999). "Antisense RNA gene therapy for studying and modulating biological processes." Cellular and Molecular Life Sciences. 55 (3): 334–58. doi:10.1007/s000180050296. PMID 10228554.
  3. ^ Beta-catenin signaling activity dissected in the early Xenopus embryo: a novel antisense approach. Heasman J, Kofron M, Wylie C. Dev. Biol. 2000. 222, 124-34
  4. ^ Kumar, Prashant; Kumar, Binod; Rajput, Roopali; Saxena, Latika; Banerjea, Akhil C.; Khanna, Madhu (November 2013). "Cross-protective effect of antisense oligonucleotide developed against the common 3' NCR of influenza A virus genome". Molecular Biotechnology. 55 (3): 203–211. doi:10.1007/s12033-013-9670-8. ISSN 1559-0305. PMID 23729285.
  5. ^ Kumar, B.; Khanna, Madhu; Kumar, P.; Sood, V.; Vyas, R.; Banerjea, A. C. (May 2012). "Nucleic acid-mediated cleavage of M1 gene of influenza A virus is significantly augmented by antisense molecules targeted to hybridize close to the cleavage site". Molecular Biotechnology. 51 (1): 27–36. doi:10.1007/s12033-011-9437-z. ISSN 1559-0305. PMID 21744034.
  6. ^ Chromatographic and related studies of alkylamide phases. B. Buszewski, P. Kasturi, R. K. Gilpin, M. E. Gangoda and M. Jaroniec, Chromatographia, August 1994, Volume 39, Issue 3–4, pages 155–161
  7. ^ Analysis of oligonucleotides by liquid chromatography with alkylamide stationary phase. Bogusław Buszewski, Zahra Safaei and Sylwia Studzińska, Open Chemistry, 2015-11-25, Volume 13, Issue 1, doi:10.1515/chem-2015-0141
  8. ^ Distler, AM; Allison, J (2001). "5-Methoxysalicylic acid and spermine: A new matrix for the matrix-assisted laser desorption/ionization mass spectrometry analysis of oligonucleotides". Journal of the American Society for Mass Spectrometry. 12 (4): 456–62. doi:10.1016/S1044-0305(01)00212-4. PMID 11322192.

Further readingEdit

  • Spingler, Bernhard (2012). "Chapter 3. Metal-Ion-Promoted Conformational Changes of Oligonucleotides". In Astrid Sigel, Helmut Sigel and Roland K. O. Sigel (ed.). Interplay between Metal Ions and Nucleic Acids. Metal Ions in Life Sciences. 10. Springer. pp. 103–118. doi:10.1007/978-94-007-2172-2_3.

External linksEdit