A guide to oligonucleotide chemistry

An oligonucleotide, often shortened to oligo, is a short single strand of DNA or RNA, typically between about twelve and fifty bases long. Where the genome in every cell runs to billions of bases, an oligo is a small, precisely defined fragment that can be designed on a screen and synthesised to order.

That precision is what makes oligos useful as medicines and as research tools. Because the sequence is written by the designer, an oligo can be programmed to recognise one specific stretch of genetic code and nothing else, acting on the instructions a cell uses rather than the proteins those instructions produce.

Every oligo is a chain of nucleotides. Each nucleotide has three parts: a sugar, a phosphate group and one of the nitrogen containing bases. In DNA the sugar is deoxyribose and the bases are adenine, cytosine, guanine and thymine. In RNA the sugar is ribose, carrying an extra hydroxyl group at the two prime position, and thymine is replaced by uracil.

Nucleotides are joined into a strand by phosphodiester bonds, which link the three prime carbon of one sugar to the five prime carbon of the next through a phosphate. This gives every strand a direction, written five prime to three prime, and a negatively charged backbone that governs how the molecule dissolves, folds and interacts with proteins.

The natural phosphodiester backbone is also the oligo's weak point: enzymes called nucleases recognise and cut it within minutes in blood or tissue. Almost every therapeutic oligo therefore carries chemical modifications, most commonly a phosphorothioate backbone, to survive long enough to work.

Two properties define an oligo. The sequence determines what it binds to, following the same base pairing rules that hold the double helix together. The chemistry, meaning the sugar, backbone and any conjugated groups, determines how stable it is, how tightly it binds, where it goes in the body and how the immune system sees it.

Designing a good oligo means balancing both at once. Syngenis One models sequence and chemistry together, screening candidates for on target activity, off target risk and manufacturability before a single base is synthesised.