Studying DNA molecules’ behavior offers insights into genetic disorders and aids in designing better drugs. Examining DNA molecules separately has been laborious and slow, and the number of possible sequence variations is massive.
Biophysicists from Delft University of Technology and Leiden University developed an innovative tool called SPARXS (Single-molecule Parallel Analysis for Rapid Exploration of Sequence space). This technique allows them to study millions of DNA molecules simultaneously, measuring millions of DNA molecules within a week instead of years or decades.
Using SPARXS, scientists gain new insights into how the structure and function of DNA depend on sequence. It can also quickly find the best sequence for applications ranging from nanotechnology to personalized medicine.
To develop the SPARXS, scientists combined two technologies: single-molecule fluorescence and next-generation Illumina sequencing. The first technique involves labeling the molecules with a fluorescent dye and visualizing them through a sensitive microscope. The latter technique reads out millions of DNA codes simultaneously.
Delft Professor Chirlmin Joo said, “It took a year to determine whether combining the two techniques is feasible, four more years to develop a working method, and two additional years to ensure accuracy and consistency in measurements while managing the vast amount of data generated.”
“The real fun and interesting part started when we needed to interpret the data. Since these experiments that combine single-molecule measurements with sequencing are completely new, we had yet to determine what results we would and could obtain. It required a lot of searching within the data to find correlations and patterns and determine the mechanisms that underly the patterns we see.”
Another challenge was handling a huge amount of data. For this, they developed an automated and robust analysis pipeline. This was hard because single molecules are delicate and produce only a tiny bit of light, which makes the data noisy. Also, the data doesn’t directly show how the DNA sequence affects its structure and movement, even with the simpler DNA we looked at.
Leiden Professor John van Noort explains, “To test our understanding, we set up a model that incorporates our knowledge of the DNA structure and compared it with the experimental data.”
Better control and understanding of DNA sequences could improve medical treatments, leading to more effective gene therapies and personalized medicine. Researchers also expect this will drive biotechnological advances and a deeper grasp of molecular biology.
Journal Reference:
- Ivo Severins, Carolien Bastiaanssen, Sung Hyun Kim, Roy B. Simons, John van Noort, Chirlmin Joo. Single-molecule structural and kinetic studies across sequence space. Science, 2024; 385 (6711): 898 DOI: 10.1126/science.adn5968