DNA Stretching: Minimizing Thermal Fluctuations for Imaging

Most microscopes have a limit to how small an object they can clearly illuminate before the smallest details start to blur together. This limitation is known as the "diffraction limit of light." However, super-resolution imaging techniques can overcome this barrier by reducing thermal fluctuations, allowing them to distinguish even the tiniest biomolecular features.

Recent research published in AIP Advances, from AIP Publishing, shows how to stretch and immobilize DNA with minimal thermal fluctuation to allow for in-depth analysis. It uses sophisticated imaging techniques and precise microfluidics control to stretch curly DNA into a straight line.

Immobilizing the molecule essentially ‘glues’ it to a substrate, preventing any movement caused by thermal fluctuations. Super-resolution imaging often requires just seconds or minutes to capture the image. During this time, thermal fluctuations random vibrations caused by the molecule’s thermal energy result in blurry images and decreased lateral resolution.”

Naoki Azuma, Study Author, Nagoya University

In the past, scientists have attempted to stretch out a DNA molecule by sticking down one end, but they discovered that the thermal fluctuations could still result in movement and blurring.

Stretching DNA refers to the process of stretching a single DNA molecule, which is originally coiled in a random coil, into a straight line. The length and structure of a single DNA molecule, its specific base sequence, and its interactions with proteins must be observed by stretching it for detailed analysis.”

Naoki Azuma, Study Author, Nagoya University

Azuma and his colleagues at Nagoya University experimented with ways to uncurl DNA molecules by applying pressure to a liquid flowing through a channel. The pressure flow provided shear force, which caused the DNA molecules to uncurl.

They discovered that regulating the liquid's flow velocity enables accurate modifications of the DNA's stretch ratio and aids in fine-tuning the shear force applied.

Accurate analysis required careful control of the stretch ratio. To “glue” the DNA molecule in place, they also employed a specialized chemical that forms chemical bonds between the DNA and a glass substrate.

While it is not yet possible to directly visualize individual base pairs, these methods enable much higher precision in observing molecular-scale structures. We aim to refine these methods to achieve higher fidelity in stretching and immobilizing DNA molecules for more accurate analysis.”

Naoki Azuma, Study Author, Nagoya University