High throughput single-molecule technology.

Abstract

Single-molecule fluorescence resonance energy transfer (smFRET) has been an indispensable tool to probe the structure of biomolecules on the nanometer scale. The intrinsic low-throughput nature of single molecule observation has, however, prevented the technique to be used to study a large pool of samples. Here, we present a highly parallel smFRET measurement scheme by combining the conventional smFRET measurement with a next generation sequencing method. A library of DNA molecules, each of which carries one of the thousands of different sequences of interest, was prepared and measured in a one pot recipe. By using this high-throughput approach, we experimentally determine the sequence-dependent end-to-end distance of single stranded DNA in various native aqueous buffer conditions.