Construction And Bioimaging Application Of DNA Nanostructures

Over the past four decades,DNA,the genetic substance in biological systems,has regarded as the basic material to construct novel nanodevices.As a nanomaterial,the remarkable advantage of DNA is manipulating target molecules with precise positioning and programming at the nanoscale;These characteristics,integrated with the biocompatibility of DNA,have promoted the potential of DNA nanostructures for materials and biological applications.One of the main goals of DNA nanotechnology is to use DNA to construct nanodevices and in turn regulate the function of biological system.The main content of this dissertation is constructing fSunTag(framework nucleic acids SunTag)nanoprobe for multicolor imaging of living cells based on DNA framework nucleic acid structures,and assembling amphiphilic DNA tetrahedron for phase transfer of hydrophobic fluorophores.Besides,we modified liposome with a density DNA shell and enhanced the signal-to-noise ratio of the hepatoma imaging.Finally,we fabricated protein-DNA functional supramolecular complexes based on the co-assembly of protein and DNA nanostructures.This work is divided into four parts and the results are summarized as follows:In chapter 2,we demonstrate the use of framework nucleic acids(FNAs)with modular design to construct FNA-SunTags(or fSunTags)for high-brightness and multiplex cellular imaging.We find that fSunTag can precisely program the copy number and spatial distribution of fluorescence molecules to construct a multicolor probe with quantized fluorescence intensity for multicolor imaging in living cells.fSunTag can also be repurposed to recruit multiple copies of aptamers for targeted imaging of cancer cells.We thus expect that fSunTag provides a generic amplification tool for quantitative and highly parallel bioimaging,bioassays and theranostics.In chapter 3,we report a simple technique for phase transfer of hydrophobic fluorophores using amphiphilic tetrahedron DNA(TDN).The amphiphilic TDN was constructed using quantitative DNA-alkyl conjugates(DNA-Alk).The incorporation of DNA-Alks in TDN assembled a hydrophobic core for weak polarity molecule carrying.The encapsulation efficiency of TDN-Alk_n is regulated by the number of alkyl chain.TDN-Alk_n are able to improve the photostability and stability against ultraviolet light photobleaching of the encapsulated fluorophores.In chapter 4,we packaged liposomes with a density DNA shell and systematically studied their metabolic profile and distribution in vivo.The DNA shell reduces the accumulation of liposomes in normal liver tissues.This material can be used as an imaging probe to distinguish liver tumor and normal liver,and to improve the signal-to-noise ratio of hepatoma imaging.In chapter 5,we fabricated a protein-DNA tetrahedron complex by co-assembly of protein and DNA tetrahedron.The protein-DNA submicrometre superstructures assembled by E.coli pilin CsgA and multi copies of DNA tetrahedral structures and maintained a linear morphology at the submicron scale.This model provides an insight into the creation of functional superstructures through the co-assembly of protein and DNA-based building blocks.In summary,we have successfully applied well-defined DNA nanostructures to living cells and biological tissues imaging through self-assembly of DNA.To further generate complexity DNA assemblies,we utilized protein molecules as templates to guide the assembly of DNA tetrahedrons forming a composite structure.