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iGEM 2012
Why make logic circuits with strand displacement? How does strand displacement work? Strand displacement reactions work in vivo!

Sensing mRNA Levels using Strand Displacement Strand Displacement NOT Gate Design Making short RNAs in vivo to use in circuits

Project Description

The complexity of engineered genetic circuits in eukaryotic systems is limited by the availability of regulatory components such as promoters, genes and repressors and is further hampered by the inability to assemble and deliver large DNA constructs. In contrast, the field of DNA computing has grown exponentially in terms of circuit complexity through use of in vitro synthetic DNA circuits that are enabled by a mechanism called toehold mediated strand displacement. These circuits have demonstrated complex digital logic with reliable and scalable behaviors in a small base-pair footprint. However, the processing power of toehold mediated strand displacement in vitro and the success of sensing dynamic inputs and actuating protein translation from traditional transcriptional and translational regulatory synthetic biology circuits have grown in parallel. Imagine, the possible adaption of strand displacement circuits into cellular environments. This could amplify the scale and complexity of biological circuits, broadening synthetic biology’s application space.

Our project leverages strand displacement to create a process technology that supports multi-input sensing, sophisticated information processing, and precisely-regulated actuation in mammalian cells. We use RNA strand displacement to sense cellular mRNA and have developed a complete logic set through strand displacement reactions by designing and testing a novel fully functioning NOT gate. Enabled by these successes and the ability to produce short RNAs in vivo, we most importantly demonstrated that toehold mediated strand displacement using RNA is a viable processing technology in vivo through demonstration of the strand displacement reporting reaction in mammalian cells. We envision in vivo RNA strand displacement as a new foundation for scaling up complexity in engineered biological systems, with applications in biosynthesis, biomedical diagnostics and therapeutics.