Inventing SBX, a DNA sequencing breakthrough

Years before, biochemist Mark Kokoris and his colleagues had begun a long quest to make the complex DNA sequencing process quicker, more accurate and more flexible.

The idea was brilliant: Overcome the challenges of directly sequencing DNA by creating a surrogate molecule with larger reporter elements. However, the task of developing the chemistry to enable these molecular elements to come together into a continuous expanded structure was technically daunting and most experts doubted the feasibility. This was an ambitious effort, and success would take Kokoris and his team on a years-long journey. 

With determination and an innovative mindset, Kokoris and his colleagues set out to create a molecule that no one had seen before. Through long hours of research, multiple failures and setbacks, but also leaps forward, the Xpandomer molecule began to take shape.

These Xpandomer molecules form the basis of breakthrough DNA sequencing technology introduced recently by Roche called sequencing by expansion (SBX). The technology has the potential to revolutionize the way DNA is sequenced, advancing scientific research for the future benefit of patients everywhere.

DNA is the body’s information storage system. It is tight and compact inside of our cells, telling the cells how to operate. For this reason, the sequencing of DNA can be important for understanding biology and also for making timely and accurate medical decisions. 

But there has always been a challenge: How can you pull information out of a very long, highly dense molecule in an efficient, flexible and cost-effective way? Technologies that have laid the foundation for the $8 billion sequencing market have all used different methods with inherent limitations. Whether needing to cycle reagents through a lengthy process or surface limitations for the reaction, there has always been a challenge to creating a technology that enables flexibility for the user to efficiently and cost effectively read the DNA code.

To enable these capabilities, Kokoris says, you must improve the signal to noise of the measurement. In other words, make it easier to distinguish the actual DNA signal from background noise, or unwanted signals. He was able to achieve this through SBX, which converts DNA information into a longer, expanded molecule - the Xpandomer - that is much easier to measure. Combine this with a nano-scale detector called a nanopore, and speed and flexibility can be achieved. An Xpandomer, with reporter codes more than 50 times longer than typical DNA bases, allows for more rapid, accurate and flexible reading when fed through the nanopore.

Kokoris, now Head of SBX Technology at Roche, started his sequencing journey in the late 1980s at the University of California, Davis. As an undergrad In the lab, he studied the molecular evolution of wildflowers and did manual Sanger sequencing work.

He was an early adopter of PCR, short for polymerase chain reaction, a common technique used today to amplify DNA segments for analysis.

After graduating, he went straight into industry, where he worked on developing a mass spec-based genotyping technology and, separately, engineering novel enzymes for gene therapy applications. He found that he could engineer proteins to do “crazy things,” and that background fed into his early thoughts about SBX. 

Kokoris spent many long days under a laboratory hood, doing experiments. The early days at Stratos Genomics, the company he co-founded with friends in 2007 to develop SBX, were rudimentary. “Funding was difficult to obtain at the time, so we had to make due with what we had,” he says. His first water bath was a pot and a hot plate, and the first oxygen-free, water-free environments were plastic bags. He reused pipette tips to save on costs. “Even though we didn’t have much to work with, it was an exciting time because there were so many possibilities in front of us. Even early on, we knew the things we wanted to do and how we wanted to evolve the technology.”

At Stratos, Kokoris continued chipping away at the SBX concept. He won investment funding and did a technical research project with the Roche Diagnostics Research & Development chemistry team from 2014-16 and again from 2018-19. After more innovation, Roche acquired Stratos in 2020, providing Roche with access to the SBX chemistry.

Innovation, you could say, is part of Kokoris’ DNA. The best mindset for innovation, he says, is unbounded thinking. You have to be like an artist, thinking about what you want to create and not limiting that creativity. Fear, he says, can fuel that creativity if used properly, just as compliance and giving in to conventional thinking can crush it.

And failure, he says, is necessary. “You have to stay disciplined and focused while continuing to push the boundaries, and realize that you can’t solve every problem at once," he says. “For every significant success we’ve had, we’ve probably had hundreds of failures.”

The lynchpin SBX experiment came after many, many failed experiments and nearly 10 years of work. 

Kokoris and team hypothesized that adding a certain molecule to the chemistry would interact with the DNA and stabilize the structure. It could be the “glue” that would hold the whole system together.

This time, they were right.

“When you innovate, you have to be willing to leave your brain open and follow the data, without worrying about being wrong,” Kokoris says. “Otherwise, you might miss the little things, like a blurry band on a gel.”