ABSTRACT
The number of applications of self-assembled deoxyribonucleic acid (DNA) origami nanoparticles (DNA NPs) has increased drastically, following the development of a variety of single-stranded template DNA (ssDNA) that can serve as the scaffold strand. In addition to viral genomes, such as M13 bacteriophage and lambda DNAs, enzymatically produced ssDNA from various template sources is rapidly gaining traction and being applied as the scaffold for DNA NP preparation. However, separating fully formed DNA NPs that have custom scaffolds from crude assembly mixes is often a multistep process of first separating the ssDNA scaffold from its enzymatic amplification process and then isolating the assembled DNA NPs from excess precursor strands. Only then is the DNA NP sample ready for downstream characterization and application. In this work, we highlight a single-step purification of custom sequence- or M13-derived scaffold-based DNA NPs using photocleavable biotin tethers. The process only requires an inexpensive ultraviolet (UV) lamp, and DNA NPs with up to 90% yield and high purity are obtained. We show the versatility of the process in separating two multihelix bundle structures and a wireframe polyhedral architecture.
Subject(s)
Biotin , DNA, Single-Stranded , Nanoparticles , Biotin/chemistry , Nanoparticles/chemistry , DNA, Single-Stranded/chemistry , DNA, Single-Stranded/isolation & purification , Bacteriophage M13/chemistry , Bacteriophage M13/genetics , DNA/chemistry , DNA/isolation & purification , Ultraviolet RaysABSTRACT
DNA nanotechnology is rapidly gaining traction in numerous applications, each bearing varying degrees of tolerance to the quality and quantity necessary for viable nanostructure function. Despite the distinct objectives of each application, they are united in their reliance on essential analytical techniques, such as purification and characterization. This tutorial aims to guide the reader through the current state of DNA nanotechnology analytical chemistry, outlining important factors to consider when designing, assembling, purifying, and characterizing a DNA nanostructure for downstream applications.
Subject(s)
Nanostructures , Nanotechnology , Nanotechnology/methods , Nanostructures/chemistry , DNA/chemistryABSTRACT
The authors report on series of side-chain smectic liquid crystal elastomer (LCE) cell scaffolds based on star block-copolymers featuring 3-arm, 4-arm, and 6-arm central nodes. A particular focus of these studies is placed on the mechanical properties of these LCEs and their impact on cell response. The introduction of diverse central nodes allows to alter and custom-modify the mechanical properties of LCE scaffolds to values on the same order of magnitude of various tissues of interest. In addition, it is continued to vary the position of the LC pendant group. The central node and the position of cholesterol pendants in the backbone of ε-CL blocks (alpha and gamma series) affect the mechanical properties as well as cell proliferation and particularly cell alignment. Cell directionality tests are presented demonstrating that several LCE scaffolds show cell attachment, proliferation, narrow orientational dispersion of cells, and highly anisotropic cell growth on the as-synthesized LCE materials.
Subject(s)
Biocompatible Materials/chemistry , Elastomers/chemistry , Liquid Crystals/chemistry , Mechanical Phenomena , Animals , Biocompatible Materials/chemical synthesis , Biocompatible Materials/pharmacology , Cell Line , Cell Movement/drug effects , Cell Proliferation/drug effects , Dermis/cytology , Elastomers/chemical synthesis , Elastomers/pharmacology , Fibroblasts/cytology , Fibroblasts/drug effects , Humans , Liquid Crystals/ultrastructure , Mice , Microscopy, Polarization , Myoblasts/cytology , Myoblasts/drug effects , Porosity , Scattering, Small Angle , Stress, Mechanical , Temperature , X-Ray DiffractionABSTRACT
3D biodegradable and highly regular foamlike cell scaffolds based on biocompatible side-chain liquid crystal elastomers have been prepared. Scaffolds with a primary porosity characterized by spatially interlaced, interconnected microchannels or an additional secondary porosity featuring interconnected microchannel networks define the novel elastomeric scaffolds. The macroscale morphology of the dual porosity 3D scaffold resembles vascular networks observed in tissue. 3D elastomer foams show four times higher cell proliferation capability compared to conventional porous templated films and within the channels guide spontaneous cell alignment enabling the possibility of tissue construct fabrication toward more clinically complex environments.
