Blog Post
Leaping ahead of Chemistry
The Path to Enzymatic DNA Synthesis
Back in June - only 4 months ago - DNA Script announced two achievements that had never before been reported: the successful enzymatic synthesis of a 50-nucleotide DNA strand and the first PCR using enzymatically synthesized primers. Since then, tremendous progress has been achieved, and enzymatic DNA synthesis is now at parity with traditional chemical technology — and poised to quickly surpass the gold standard in speed, efficacy and length.

The purpose of this series of blog posts is to demonstrate the capabilities of DNA Script's technology and how this technology will serve as a foundation for easily programmable biology. Most of the results shown here have been presented by our CEO, Thomas Ybert at SynBioBeta 2018. This series will be divided into 3 separate parts, published over the next 2 weeks:

  1. Enzymatic DNA synthesis performance
  2. Applications: DNA data storage & Epigenetics
  3. The path to an easily programmable biology

Part 1 - Enzymatic DNA synthesis performance

The potential of enzymatic DNA synthesis has been recognized for many years : enzymes such as polymerases are extremely efficient tools for building DNA. The challenge we faced when building this technology was mastering every aspect in the synthesis process so that the efficiency could be as high as possible. Proving the technology works on long strands of DNA is the only way to be sure that our technology can match, and eventually surpass, existing chemical approaches.

We are very proud to announce that DNA Script's technology can now perform as well or better than phosphoramidite chemistry technology. The table below sums up how DNA Script's approach stacks up:

The cycle efficacy is the most critical criteria when looking at DNA synthesis performance. The best yield for chemical synthesis is 99.5% per cycle, but most industrial facilities are between 98.5% and 99.3%. The Synthesis Team at DNA Script successfully attained 99.5% average cycle efficacy on synthesis of 150 nucleotides long oligonucleotides. Even higher performance on shorter strands/sequences were reached.
Fig. 1 - This figure shows the result of an electrophoresis analysis of synthesis of an oligonucleotide with 100 nucleotides in length. More than 58% of the DNA fragments successfully reached 100 nucleotides, which makes a 99.5% average efficacy per cycle - the gold standard for chemical synthesis. Each cycle was performed in 5 minutes, on a fully automated platform without any gas or humidity control.
Reducing cycle time is very useful for increasing throughput of an instrument or factory, and is critical for certain applications where DNA is a key reagent whose turnaround time is a limiting factor. Synthetic Biology should be like software programming; no one should be waiting several weeks for DNA constructs and the field needs to iterate experiments as quickly as possible. Today, adding one nucleotide takes 5 minutes in routine with our technology, compared to an average 10 minutes for traditional synthesis. We hope to be able to add each nucleotide in under 3 minutes by next year.

The maximum length synthesized is a key parameter as increasing it can open new market segments. Chemical technology ranges between 150nt and 200nt in length, the longer fragments being much more expensive as they require special care. Last month, DNA Script managed to synthesize a 150nt strand in one shot and proved that our technology can scale. We are very confident we can further increase length.

One of the biggest differentiators of enzymatic DNA synthesis is that it is performed in water-based media. This avoids many problems (organic solvent sourcing and waste management, solvents harming DNA, branching, argon chambers, etc) and opens a new world of capabilities - for instance developing a benchtop DNA printer.

The performance we have achieved can be reached with the 4 natural nucleotides (and even more than those 4 nucleotides, read next week's post!) with very similar efficacy.

Our team's main focus is now to expand what we call the "sequence space" - meaning all the sequences that can be synthesized successfully to meet customers' needs. Every technology has some bias: phosphoramidite chemistry struggles to synthesize and purify sequences with a high GC content. Assessing and solving the biases of our technology is a key priority for us, which is why we do Next Generation Sequencing on our fragments. It is important to note that we do not purify our DNA whatsoever before sequencing: the result below is from a raw synthesis, straight from the tube. The full sequence correctness is
Fig. 2 - This figure shows the result of a NGS analysis of a 50nt. The percentage of 100% correct sequence was 60% for this sequence. The "Identity" color and heights represents percentage of sequence correctness at each position. Any error of more than 3% on any position would show in red.
The raw performance of DNA Script's enzymatic DNA synthesis is now at parity with the best existing commercial technology.
More than results, what is really thrilling to the team is the pace of our development which far outperforms the pace of chemical DNA synthesis introduced close to 50 years ago.

Fig. 3 - The graph above shows the the max length achieved by chemical and enzymatic technology since inception. The Y axis is in log scale.
The potential moving forward and the impact it will have on SynBio is overwhelming.

The next week post will present results and proofs of concept on applications we started working on. Stay tuned!
OCT, 2nd 2018