Single-Molecule Methods for Characterizing Different DNA Higher-Order Structures

DNA is considered to be not only a carrier of the genetic information of life but also a highly programmable and self-assembled nanomaterial. Different DNA structures are related to their biological and chemical functions. Hence, understanding the physical and chemical properties of various DNA structures is of great importance in biology and nanochemistry. However, the bulk assay ignores the heterogeneity of DNA structures in solution. Single-molecule methods are powerful tools for observing the behavior of individual molecules and probing the high heterogeneity of free energy states. In this review, we introduce single-molecule methods, including single-molecule detection and manipulation methods, and discuss how these methods can be conducive to measuring the molecular properties of single-/double-stranded DNA (ss/dsDNA), DNA higher-order structures, and DNA nanostructures. We conclude by providing a new perspective on the combination of DNA nanotechnology and single-molecule methods to understand the biophysical properties of DNA and other bio-matter and soft matter.