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Meta-DNA: A DNA-Based Approach to Synthetic Biology

Nikhil Gopalkrishnan
Bernard Yurke
John H. Reif
Systems and Synthetic Biology: A Systematic Approach (2014)


The goal of synthetic biology is to design and assemble synthetic systems that mimic biological systems. One of the most fundamental challenges in synthetic biology is to synthesize artificial biochemical systems, which we will call meta-biochemical systems, that provide the same functionality as biological nucleic acids-enzyme systems, but that use a very limited number of types of molecules. The motivation for developing such synthetic biology systems is to enable a better understanding of the basic processes of natural biology, and also to enable re-engineering and programmability of synthetic versions of biological systems. One of the key aspects of modern nucleic acid biochemistry is its extensive use of protein enzymes that were originally evolved in cells to manipulate nucleic acids, and then later adapted by man for laboratory use. This practice provided powerful tools for manipulating nucleic acids, but also limited the extent of the programmability of the available chemistry for manipulating nucleic acids, since it is very difficult to predictively modify the behavior of protein enzymes. Meta-biochemical systems offer the possible advantage of being far easier to re-engineer and program for desired functionality. The approach taken here is to develop a biochemical system which we call meta-DNA (abbreviated as mDNA),Meta-DNA (mDNA) based entirely on strands of DNA as the only component molecules. Our work leverages prior work on the development of self-assembled DNA nanostructures. Each base of a mDNA Meta-DNA (mDNA)is a DNA nanostructure. Our mDNA bases are paired similar to DNA bases, but have a much larger alphabet of bases, thereby providing increased power of base addressability. Our mDNA bases can be assembled to form flexible linear assemblies (single stranded mDNA) analogous to single stranded DNA, and can be hybridized to form stiff helical structures (duplex mDNA) analogous to double Double strand meta-DNA (dsmDNA) stranded DNA, and also can be denatured back to single stranded mDNA. Our work also leverages the abstract activatable tile model developed by Majumder et al. and prior work on the development of enzyme-free isothermal protocols based on DNA hybridization and sophisticated strand displacement hybridization reactions. We describe various isothermal hybridization reactions that manipulate our mDNA in powerful ways analogous to DNA–DNA reactions and the action of various enzymes on DNA. These operations on mDNA include (i) hybridization of single strand mDNA (ssmDNA)Single strand meta-DNA (ssmDNA) into a double strand mDNA (dsmDNA)Double strand meta-DNA (dsmDNA) and heat denaturation of a dsmDNA Double strand meta-DNA (dsmDNA)into its component ssmDNA Single strand meta-DNA (ssmDNA)(analogous to DNA hybridization and denaturation), (ii) strand displacement of one ssmDNA Single strand meta-DNA (ssmDNA)by another (similar to strand displacement in DNA), (iii) restriction cuts on the backbones of ssmDNA Single strand meta-DNA (ssmDNA)and dsmDNA Double strand meta-DNA (dsmDNA)(similar to the action of restriction enzymes on DNA), (iv) polymerization chain reactions that extend ssmDNA Single strand meta-DNA (ssmDNA)on a template to form a complete dsmDNA Double strand meta-DNA (dsmDNA)(similar to the action of polymerase enzyme on DNA), (v) isothermal denaturation of a dsmDNA Double strand meta-DNA (dsmDNA)into its component ssmDNA Single strand meta-DNA (ssmDNA)(like the activity of helicase enzyme on DNA) and (vi) an isothermal replicator reaction which exponentially amplifies ssmDNA Single strand meta-DNA (ssmDNA)strands (similar to the isothermal PCR reaction). We provide a formal model to describe the required properties and operations of our mDNA, and show that our proposed DNA nanostructures and hybridization reactions provide these properties and functionality.

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