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iGEM 2012

Background and Motivation

In the near future, biological circuits will be much more modular and sophisticated than they are now, with a ten-fold smaller nucleotide footprint.

The Enabling Technology: Toehold-Mediated Strand Displacement

A gate strand and output strand exist as a complex that is partially bound through complementary Watson-Crick base-pairing within the S2 binding domain. The gate strand also contains an open, unbound domain called a toehold region, T*. An input strand with a free complementary toehold region, T, can bind to the toehold region on the gate strand, and subsequently displace the output strand to yield an input-gate complex. This is called a toehold-mediated strand displacement reaction. The output strand is used as an input for a downstream gate-output complex.


Qian and Winfree (Science 2011) utilized DNA computation to create AND and OR logic gates in vitro. They constructed a sophisticated binary square root circuit using these gates:
Each of these:

undergoes the same toehold-mediated strand displacement reaction. These reactions are fully modular and can be scaled to circuits of any degree of sophistication.

Motivation for Bringing Strand Displacement to Mammalian Synthetic Biology

  • More sophisticated circuits with smaller nucleotide footprint
    A traditional NOT gate is ~1000 bp, whereas our strand displacement NOT gate is ~ gate is ~100 bp.
  • Simple combinatorial design space
    With 4 bases, we can create a nearly infinite number of orthogonal sequences leading to orthogonal parts.
  • Ease of composition
    The input motif matches the output motif allowing for modular cascading reactions.
  • Tunability
    We can set arbitrary digital signal thresholds by varying the concentration of circuit species. We can also achieve signal amplification by including a fuel molecule.