Abstract
Magnetic tweezers is a versatile yet simple single-molecule manipulation technique that has been used to study a broad range of nucleic acids and nucleic acid-based molecular motors. In this chapter, we combine micro-mirror-based total internal reflection microscopy with a magnetic tweezers instrument, permitting simultaneous single-molecule visualization and mechanical manipulation. We provide a simple method to calibrate the evanescent wave penetration depth via supercoiling of DNA with a fluorescent nanodiamond-labeled magnetic bead and a complementary method employing a surface-immobilized fluorescent nanodiamond.