To our knowledge, we have developed a novel temperature-jump optical tweezers setup that changes the temperature locally and rapidly. It uses a heating laser with a wavelength that is highly absorbed by water so it can cover a broad range of temperatures. This instrument can record several force-distance curves for one individual molecule at various temperatures with good thermal and mechanical stability. Our design has features to reduce convection and baseline shifts, which have troubled previous heating-laser instruments. As proof of accuracy, we used the instrument to carry out DNA unzipping experiments in which we derived the average basepair free energy, entropy, and enthalpy of formation of the DNA duplex in a range of temperatures between 5°C and 50°C. We also used the instrument to characterize the temperature-dependent elasticity of single-stranded DNA (ssDNA), where we find a significant condensation plateau at low force and low temperature. Oddly, the persistence length of ssDNA measured at high force seems to increase with temperature, contrary to simple entropic models.
A main goal of single-molecule experiments is to evaluate equilibrium free energy differences by applying fluctuation relations to repeated work measurements along irreversible processes. We quantify the error that is made in a free energy estimate by means of the Jarzynski equality when the accumulated work expended on the whole system (including the instrument) is erroneously replaced by the work transferred to the subsystem consisting of the sole molecular construct. We find that the error may be as large as 100%, depending on the number of experiments and on the bandwidth of the data acquisition apparatus. Our theoretical estimate is validated by numerical simulations and pulling experiments on DNA hairpins using optical tweezers.
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