Stretching submicron biomolecules with constant-force axial optical tweezers

Optical tweezers have become powerful tools to manipulate biomolecular systems, but are increasingly difficult to use when the size of the molecules is <1 μm. Many important biological structures and processes, however, occur on the submicron length scale. Therefore, we developed and characterized an optical manipulation protocol that makes this length scale accessible by stretching the molecule in the axial direction of the laser beam, thus avoiding limiting artifacts from steric hindrances from the microscope coverslip and other surface effects. The molecule is held under constant mechanical tension by a combination of optical gradient forces and backscattering forces, eliminating the need for electronic feedback. We demonstrate the utility of this method through a measurement of the force-extension relationship of a 1298 bp ds-DNA molecule.