Design of Novel Magnetic Tweezers and its use for Studying DNA-Compacting Proteins

We report the development of a horizontal magnetic tweezers capable of applying forces in the 0.1 - 100 piconewton (pN) range on single DNA molecules. The two ends of the DNA molecule are attached to 2.8 um paramagnetic beads with one bead immobilized on a rigid glass surface while the other bead is suspended near a small bar magnet. The magnet is moved towards the tethered DNA at a speed between 0.25 – 10 um/s and can generate forces greater than 70 pN on the DNA molecules. These forces are produced in the focal plane of the microscope objective, permitting direct DNA extension measurements with a precision of <10 nm. We calibrate the tweezers using the DNA overstretching transition, and study hysteretic effects in extension-compaction cycles. We describe the tweezers in detail and present data validating its performance. We conclude with a discussion of our ongoing single experiments on the binding mechanism of the protein mIHF, a protein that may play an important role in the infection pathway of tuberculosis.