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Easy and inexpensive method for linking other molecules to DNA sequences with desired functions

The development of new types of drugs based on DNA and RNA is happening at a rapid pace. The mRNA vaccines against COVID-19 are just one of many examples of DNA-based drugs on the market. Many are on the way, but the production of the necessary DNA is costly. A research group at Aarhus University has now found an effective and inexpensive method for producing stable DNA sequences attached to chemical groups that e.g., ensures that the drugs are directed into the right cells.

Professor Kurt Gothelf and Ph.D. Angel Santorelli. (Photo: Aarhus University)
Professor Kurt Gothelf and Ph.D. Angel Santorelli has published new results in the journal, Nucleic Acids Research, which can streamline and reduce the costs of DNA production. (Photo: Aarhus University)
DNA helix. Illustr.: Colourbox
Researchers from Aarhus University have developed a method to easily and cheaply produce desired modified DNA sequences. (Illustration: Colourbox)
Illustration of DNA helix with chemical modification. (Nucleic Acids Research, 2022, gkac566)
Positioning of a chemical modification in a DNA helix, which is introduced using new and inexpensive method. (Illustration: Nucleic Acids Research, 2022, gkac566)

The production of short DNA and RNA sequences, called oligonucleotides, is today essential in connection with diagnostics (COVID-19 test), DNA sequencing, gene synthesis for synthetic biology and gene therapy, such as in the CRISPR-Cas9 method.

The technology of oligonucleotide production has been advanced since it was developed in 1981, but the underlying chemistry remains largely unchanged. One is still dependent on phosphoramidites, which are chemical compounds that, unfortunately, are unstable in the production process.

However, Kurt Gothelf and his research group have previously shown a more efficient way to produce the building blocks for oligonucleotides by developing a flow-based method to make phosphoramidites immediately before they are to be used, thus avoiding this degradation. You can read more about this in a previous article here: New method for producing synthetic DNA.

Expensive to acquire DNA sequences with special functionalities

There is a great need for oligonucleotides that have been modified with special functionalities. For example, oligonucleotides modified with certain sugars have been shown to be taken up by the liver. Based on this, several oligonucleotide drugs are used to regulate proteins that are, in fact, produced in the liver and thus can cure diseases. It is also important to be able to attach dyes to oligonucleotides as this is often used in diagnostics and research.

Usually, these functionalities are inserted into oligonucleotides by the building blocks, the phosphoramidites, being produced with a specific chemical handle or the desired functional group. There are commercial catalogs with several hundred varieties of these modified phosphoramidites. However, such modified phosphoramidites are, in most cases, very expensive to obtain and, as mentioned above, they are unstable in liquids at room temperature.

New easy and cheap method for synthesis of special building blocks

In a recently published scientific article published in Nucleic Acids Research, which is the leading oligonucleotide journal, Angel Santorelli and Kurt Gothelf present an easy and inexpensive method of introducing modifications to oligonucleotides without the need for special phosphoramidites.

For this purpose, they use chemical compounds called sulfonyl azides, which can provide oligonucleotides with many different functionalities during the chemical synthesis of the oligonucleotides.

The method is compatible with the previously mentioned automated and flow-based method for oligonucleotide synthesis, where degradation of the phosphoramidites is avoided. The recently published method, for the synthesis of modified phosphoramidites, can thus serve as a simple alternative to the unstable and expensive special phosphoramidites currently used in the production of modified oligonucleotides.


SUPPLEMENTARY INFORMATION, INCLUDING CONTACT INFORMATION

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External funding

European Commission (DNA Robotics, 765703)
Novo Nordisk Foundation (CEMBID, NNF17OC0028070)

Conflicts of interest

The researchers declare that there are no conflicts of interest.

Link to scientific paper

Conjugation of chemical handles and functional moieties to DNA during solid phase synthesis with sulfonyl azides.
Angel Santorelli and Kurt Gothelf. Nucleic Acids Research. 2022, gkac566
https://academic.oup.com/nar/advance-article/doi/10.1093/nar/gkac566/6633897?login=true

Contact information

Professor Kurt Gothelf
Coordinator of the Marie Skłodowska-Curie ITN, DNA Robotics
Director of Center for Multifunctional Biomolecular Drug Design
Interdisciplinary Nanoscience Center (iNANO) & Department of Chemistry
Aarhus University
Email: kvg@chem.au.dk