The ability to manipulate and probe biomolecules at the single-molecule level has become an essential approach for understanding molecular interactions, conformational dynamics, and nanoscale transport phenomena. Advances in experimental techniques have enabled precise control of individual molecules with high spatial resolution and piconewton-level force sensitivity. These developments have significantly expanded the capability of studying biomolecular mechanics and dynamics beyond conventional ensemble measurements. A variety of physical strategies have been developed for single-molecule manipulation, including mechanical-force-based approaches, electric-field-driven methods, and nanoscale structural confinement techniques. Mechanical-force-based methods, such as optical tweezers, magnetic tweezers, and atomic force microscopy, enable direct measurement of molecular mechanical responses. Electric-field-based manipulation, represented by dielectrophoresis, allows noncontact control of particles and biomolecules through polarization effects in non-uniform electric fields. In addition, nanopore-based systems employ nanoscale confinement to regulate molecular transport and residence behavior. This review provides an overview of representative single-molecule manipulation techniques based on mechanical, electrical, and structural control and discusses their fundamental principles and implementation strategies.