Droplet systems, such as DNA droplets, formed by the liquid-liquid phase separation of macromolecules, play an important role in cellular functions. Computational DNA droplets, which can recognize specific patterns in tumor biomarker microRNA sequences, have now been developed by combining the technologies of DNA droplets and DNA computing.
Aqueous droplet formation in macromolecules via liquid-liquid phase separation (or coacervation) is a hot topic in life sciences research. DNA is one of the most interesting macromolecules that form droplets because it is predictable and programmable, both of which are useful in nanotechnology. The programmability of DNA has recently been used to construct and regulate DNA droplets formed by coacervation of sequence designed DNAs.
A group of scientists at Tokyo University of Technology (Tokyo Tech) led by Prof. Masahiro Takinoue has developed a computational DNA droplet with the ability to recognize specific combinations of chemically synthesized microRNAs (miRNAs) that act as biomarkers of tumors. Using these miRNAs as molecular input, the droplets can give a DNA logic computing output through physical DNA droplet phase separation.
If a DNA droplet that can integrate and process multiple inputs and outputs can be developed, it could be used in early disease detection as well as drug delivery systems. Our current study also serves as a springboard for future research into intelligent artificial cells and molecular robots.
Prof. Masahiro Takinoue
Prof. Takinoue explains the need for such studies, “The applications of DNA droplets have been reported in cell-inspired microcompartments. Even though biological systems regulate their functions by combining biosensing with molecular logical computation, no literature is available on integration of DNA droplet with molecular computing.” Their findings were published in Advanced Functional Materials.
A series of experiments were required to create this DNA droplet. To begin, they created three types of Y-shaped DNA nanostructures known as Y-motifs A, B, and C with three sticky ends to create A, B, and C DNA droplets. Similar droplets typically band together automatically, whereas dissimilar droplets require a special “linker” molecule. To connect the A droplet to the B and C droplets, they used linker molecules known as AB and AC linkers, respectively.
They tested the “AND” operation in the AB droplet mixture in their first experiment by introducing two input DNAs. The presence of input is recorded as 1 in this operation, while its absence is recorded as 0. The phase separation of the AB droplet mixture occurred only at (1,1), indicating that the AND operation was successfully applied. Following the findings of this study, the researchers decided to add breast cancer tumor markers, miRNA-1 and miRNA-2, to an AC droplet mixture as inputs for the AND operation. The AND operation was successful, indicating that the miRNAs were identified by the computational DNA droplet.
In subsequent experiments, the researchers demonstrated simultaneous AND and NOT operations in an AB mixture containing the breast cancer biomarkers miRNA-3 and miRNA-4. Finally, they made an ABC droplet mixture and added all four breast cancer biomarkers to it. The phase separation in an ABC droplet was determined by the cleavage of the linker, which resulted in a two-phase separation or a three-phase separation.
This property of the ABC droplet allowed the researchers to demonstrate the ability to detect a set of known cancer biomarkers or markers of three diseases at the same time. Prof. Takinoue, who is also the corresponding author, believes computational DNA droplets have enormous potential. He claims that “If a DNA droplet that can integrate and process multiple inputs and outputs can be developed, it could be used in early disease detection as well as drug delivery systems. Our current study also serves as a springboard for future research into intelligent artificial cells and molecular robots.”
