ENGINEERING DNA-BASED NANOCHANNELS AND VESICLES FOR CONTROLLED MOLECULAR TRANSPORT
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For the past two decades, synthetic transmembrane nanopores and nanochannels have become powerful tools in biosensing and single-molecule studies. Due to the ease of rational design and advancements in DNA functionalization, DNA has been established to be versatile building blocks for the bottom-up fabrication of nanostructures. Recently, DNA-based nanopores in both small diameters (1-2 nm) showing transport of ions and small molecules and large diameters (5-10 nm) showing transport of proteins across lipid bilayer membranes were reported. Nevertheless, those DNA nanopores have lengths below 100 nm, and the molecular transport only occurs across lipid membranes. It remains unknown if longer nanochannels can be constructed for transport over extended distances. Such nanochannels of longer lengths can be potentially used as conduits for carrying molecules on the cell-size scale or between compartments apart. We have designed a microns-long DNA nanochannel 7 nm inner diameter that inserts onto the lipid membranes of giant unilamellar vesicles and allows the transport of small molecules through its barrel. Kinetics analysis suggests a continuum diffusion model can describe the transport phenomenon within the DNA nanochannel. The reduced transport upon bindings of DNA origami caps to the channel ends reveals the molecules mainly transport from one channel end to the other rather than leak across channel walls. We further design a DNA nanopore-cap system that responds to specific DNA sequences. In combination with giant unilamellar vesicles that encapsulate glucose molecules, we present a biosensor system consisting of capped DNA nanopores and vesicles that can detect and amplify nanomolar DNA signals millimolar glucose outputs. The DNA-based biosensor we developed shows the potentials to be used as point-of-care nucleic acid diagnostic devices. Another challenge in using DNA nanochannels or other DNA-based nanostructures in biological environments or cell culture is that they may be degraded by enzymes found in these environments, such as nucleases. To improve the DNA nanostructures' stability, we demonstrate a means by which degradation can be reversed in situ through the repair of nanostructure defects. The ability to repair nanostructures, such as DNA nanochannels, could allow particular structures or devices to operate for long periods of time and might offer a single means to resist different types of chemical degradation.