Abstract
Nanopores can probe the structure of biopolymers in solution; however, diffusion makes it difficult to study the same molecule for extended periods. Here we report devices that entropically trap single DNA molecules in a 6.2-femtolitre cage near a solid-state nanopore. We electrophoretically inject DNA molecules into the cage through the nanopore, pause for preset times and then drive the DNA back out through the nanopore. The saturating recapture time and high recapture probability after long pauses, their agreement with a convection-diffusion model and the observation of trapped DNA under fluorescence microscopy all confirm that the cage stably traps DNA. Meanwhile, the cages have 200 nm openings that make them permeable to small molecules, like the restriction endonuclease we use to sequence-specifically cut trapped DNA into fragments whose number and sizes are analysed upon exiting through the nanopore. Entropic cages thus serve as reactors for chemically modifying single DNA molecules.
Highlights
Nanopores can probe the structure of biopolymers in solution; diffusion makes it difficult to study the same molecule for extended periods
This presents a serious epistemic challenge, as nanopore sensors should ideally discover the characteristics of a molecule without prior knowledge
Rant et al.[24] took an important step towards that goal when they developed devices comprising two pores and a microscale cavity. They demonstrated that the diffusion of particles and DNA can be significantly slowed inside the cavity[25]. Those devices held DNA molecules in the cavity for only a few seconds, which is too short to be practical for studying chemical interactions. (In a separate set of experiments, similar devices were used to measure the transit time of DNA molecules moving between two opposing nanopores by electrophoresis26)
Summary
Nanopores can probe the structure of biopolymers in solution; diffusion makes it difficult to study the same molecule for extended periods. Oligonucleotide hybridization probes[23] bind to specific DNA target sequences and create bulges that are easy to detect when they pass through a nanopore Applying this principle, Singer et al.[14] mapped the locations of target sequences on long, doublestranded DNA molecules, and Wanunu et al.[12] identified different microRNAs. To correctly infer that the structure of a DNA molecule has changed, it is crucial to know what was the structure of that molecule before the chemical interaction occurred. Entropic cages are useful as single-molecule chemical reactors for nanopore biosensing with a variety of chemical probes[29,30], and for studying biologically significant interactions between DNA and other molecules
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