Since 1953, when the double helix structure of DNA was first proposed by James Watson and Francis Crick, several seminal technological breakthroughs have led to the emergence of the field that we now call biotechnology. Due to the fact that the qualities and capabilities of all organisms are encoded in their genomes, encoded in the four letters of the nucleic acids DNA and RNA, many of these advances were related to deciphering DNA sequence, creating or manipulating DNA and RNA sequences either in a test tube or in an organism and establishing the connection between sequences encoded in the genome and traits or qualities of the organism. Arguably the most important technology for future developments in biotechnology, and especially in the field of synthetic biology that concerns itself with creating microorganisms and proteins for myriad 21st century applications and uses, is the ability to synthesize DNA from scratch.

Synthesizing a new DNA sequence has been possible since the 1980s, and synthetic DNA has found many applications in R&D and industry. The volume of synthesized DNA has increased exponentially, driven in recent years by the expansion of nucleic acid therapeutics, vaccines and diagnostics, the mRNA-based vaccines against COVID-19 being a particularly well-known example, but far from the only one.

Although the speed and cost of DNA and RNA synthesis have dropped as improvements have been made to standard synthesis methods, the decrease in cost has been relatively modest compared to DNA sequencing which is now multiple orders of magnitude faster and cheaper since the first sequencing system was developed in by Allan Maxam and Walter Gilbert in 1976. The fact is that DNA and RNA remain relatively expensive compared to what they cost in the 1980s and 1990s, and this is holding back progress in biotechnology and medicine.

So how is DNA synthesized for pharmaceutical, industrial and R&D uses?

The answer is both simple and strange, because the established process for de novo DNA synthesis uses synthetic chemistry, which offers flexible and efficient methods for addition of one nucleotide, or DNA/RNA building block, at a time to a growing strand.

Why is this strange?

Because DNA is a biological molecule, and the world of biology has evolved countless and innumerable ways of synthesizing DNA and RNA in cells, in the form of enzymes (nature’s catalysts), specifically DNA and RNA polymerases. And in the past, when man has had a need for a technology or solution related to biology, these have always been readily discoverable in nature.

So if nature is full of nucleic acid polymerases capable of synthesizing DNA and RNA, why do we not use these industrially? Why do we have to invent and use our own chemical methods that are both less efficient and more expensive?

The answer lies in the manner in which DNA synthesis is practiced naturally. Nature is not a Shakespeare, Agatha Christie or Hemingway who can conjure up words, sentences and plots from figments of their imagination. Rather, nature is more like a medieval scribe, copying a manuscript by candlelight, letter by letter, occasionally introducing an error or changing its meaning. In nature, DNA and RNA are copied from other DNA and RNA molecules, and only rarely are nucleic acid sequences synthesized without a template (in molecular biology jargon, in a template-independent manner). By this quirk of biology and its extreme reliance on existing sequences as the basis for synthesizing new ones, the vast majority of DNA and RNA polymerases in nature require an existing strand of DNA or RNA to create a new one. For the Jane Austens, Cervantes’ and Steinbecks of the genetic world, nature is surprisingly poor in tools that allow them to write entirely new genetic sequences.

Which is how synthetic chemistry stepped in to fill a void. In the 1980s, when the first scientists tried to devise methods to synthesize DNA, the world of chemistry seemed to offer the better practical solutions. Marvin Caruthers and his co-workers developed chemically activated nucleotides called phosphoramidites. Hiding behind this tongue-twister is a highly effective method for joining nucleotides that is still widely practiced today and used to synthesize the vast majority of man-made DNA. But after decades of optimization, the phosphoramidite method has reached its limits in terms of synthesis cost, while retaining its drawbacks which include difficulty in synthesizing at large scales and a requirement of toxic organic solvents, which complicate the expansion of DNA and RNA synthesis capacity.

The limitations of the phosphoramidite method have motivated scientists to take another, closer look at the use of enzymes for industrial DNA and RNA synthesis. Starting about a decade ago, enzymologists imagined that evolving specialized, template-independent DNA polymerases may allow the development of an enzymatic DNA synthesis process. This has led multiple companies, DNAScript, Ansa Biotechnologies, Molecular Assemblies and Primordial Genetics among them, to perfect nucleic acid polymerases for the template independent addition of single nucleotides. DNAScript is starting to market an oligonucleotide synthesis system for laboratory use. The other companies are not far behind in readying their own processes. Will they succeed in removing the nucleic acid synthesis bottleneck to enable, inexpensive, rapid, scalable and environmentally friendly synthesis of nucleic acids?