Shogo Hamada

Construction of a genetic AND gate under a new standard for assembly of genetic parts

BackgroundAppropriate regulation of respective gene expressions is a bottleneck for the realization of artificial biological systems inside living cells. The modification of several promoter sequences is required to achieve appropriate regulation of the systems. However, a time-consuming process is required for the insertion of an operator, a binding site of a protein for gene expression, to the gene regulatory region of a plasmid. Thus, a standardized method for integrating operator sequences to the regulatory region of a plasmid is required.ResultsWe developed a standardized method for integrating operator sequences to the regulatory region of a plasmid and constructed a synthetic promoter that functions as a genetic AND gate. By standardizing the regulatory region of a plasmid and the operator parts, we established a platform for modular assembly of the operator parts. Moreover, by assembling two different operator parts on the regulatory region, we constructed a regulatory device with an AND gate function.ConclusionsWe implemented a new standard to assemble operator parts for construction of functional genetic logic gates. The logic gates at the molecular scale have important implications for reprogramming cellular behavior.

Quantitative analysis of molecular-level DNA crystal growth on a 2D surface

Crystallization is an essential process for understanding a molecules aggregation behavior. It provides basic information on crystals, including their nucleation and growth processes. Deoxyribonucleic acid (DNA) has become an interesting building material because of its remarkable properties for constructing various shapes of submicron-scale DNA crystals by self-assembly. The recently developed substrate-assisted growth (SAG) method produces fully covered DNA crystals on various substrates using electrostatic interactions and provides an opportunity to observe the overall crystallization process. In this study, we investigated quantitative analysis of molecular-level DNA crystallization using the SAG method. Coverage and crystal size distribution were studied by controlling the external parameters such as monomer concentration, annealing temperature, and annealing time. Rearrangement during crystallization was also discussed. We expect that our study will provide overall picture of the fabrication process of DNA crystals on the charged substrate and promote practical applications of DNA crystals in science and technology.

Size‐Controllable DNA Rings with Copper‐Ion Modification

by simple molecular modi-fication, it makes DNA nanostructures one of the most viable biomaterials for use with current techniques. Although many developments took advantage of these characteristics, what has been lacking in DNA nanotechnology is sufficient investi-gation into specific interactions between DNA nanostructures and metal ions. Due to DNA’s poor conductivity,

Self-replication of DNA rings

Biology provides numerous examples of self-replicating machines, but artificially engineering such complex systems remains a formidable challenge. In particular, although simple artificial self-replicating systems including wooden blocks, magnetic systems, modular robots and synthetic molecular systems have been devised, such kinematic self-replicators are rare compared with examples of theoretical cellular self-replication. One of the principal reasons for this is the amount of complexity that arises when you try to incorporate self-replication into a physical medium. In this regard, DNA is a prime candidate material for constructing self-replicating systems due to its ability to self-assemble through molecular recognition. Here, we show that DNA T-motifs, which self-assemble into ring structures, can be designed to self-replicate through toehold-mediated strand displacement reactions. The inherent design of these rings allows the population dynamics of the systems to be controlled. We also analyse the replication scheme within a universal framework of self-replication and derive a quantitative metric of the self-replicability of the rings.

Theoretical model of substrate-assisted self-assembly of DNA nanostructures

Substrate-assisted self-assembly [S. Hamada and S. Murata, Angew. Chem., Int. Ed., 2009, 48, 6820; X. Sun et al., J. Am. Chem. Soc., 2009, 131, 13 248] is a novel methodology for DNA self-assembly to fabricate large-scale DNA nanostructures on substrate surfaces. Although a qualitative explanation of this phenomenon and some experimental results exist, the mechanism is not yet thoroughly understood. A theoretical framework will improve understanding and enable us to exploit this phenomenon for further development and future applications. Here, we propose a model and simulations describing this phenomenon, with comparison to experimental results. The model is based on simple thermodynamics and was converted into quasi-static kinetics to trace the overall process of adsorption and self-assembly during annealing. As an example, a simulation of T-motif Ring formation based on this model successfully reproduced the difference between yields in solution and on a surface, consistent with experimental observations.

Polymorphic Ring-Shaped Molecular Clusters Made of Shape-Variable Building Blocks

Self-assembling molecular building blocks able to dynamically change their shapes, is a concept that would offer a route to reconfigurable systems. Although simulation studies predict novel properties useful for applications in diverse fields, such kinds of building blocks, have not been implemented thus far with molecules. Here, we report shape-variable building blocks fabricated by DNA self-assembly. Blocks are movable enough to undergo shape transitions along geometrical ranges. Blocks connect to each other and assemble into polymorphic ring-shaped clusters via the stacking of DNA blunt-ends. Reconfiguration of the polymorphic clusters is achieved by the surface diffusion on mica substrate in response to a monovalent salt concentration. This work could inspire novel reconfigurable self-assembling systems for applications in molecular robotics.

Extending the Geometrical Design of DNA Nanostructures

Structural DNA nanotechnology enables us to design and fabricate shapes and patterns at nanoscale as a versatile platform for nanotechnology and bio-related computing. Since the introduction of crossover junctions, an endeavor to create nanostructures by DNA are now flourished as self-assemblies of various 2-D and 3-D shapes. Those achievements mainly owe to two factors: one is the geometry defined by crossover junctions, and the other is the introduction of design approach. The design approach itself is not dependent on any junction structure, however the lack of choice in junctions limits the appearance of resultant nanostructures. We found our interconnected single-duplex DNA junction extends the geometry of DNA nanostructures into a broader class of shapes and patterns. Here we propose an abstraction method that enables us to design variety of structures by those junctions with compatibility. Several demonstrations by this abstraction and possibilities of various new shapes and patterns based on the design approach are presented.