Progress toward construction and modelling of a tri-stable toggle switch in E. coli

In 2000, Gardner and Collins reported the construction of a fundamental genetic regulatory device, the bi-stable toggle switch, in E. coli. We report here our work on a natural extension of this powerful device, a tri-stable genetic toggle switch capable of switching among three stable states. Like the bi-stable switch, the tri-stable switch consists of repressible promoters that produce inhibitory proteins and requires only a transient pulse of chemical inducer to switch among stable states. Our proof-of-principal construct is designed to control the expression of three different fluorescent reporters using the pBad/AraC, pLacI/LacI, and pTetR/TetR systems; though a tri-stable switch can theoretically be constructed from any three repressible promoters that satisfy a certain mathematical relationship. We have modelled the system extensively, creating both a simple continuous deterministic model based on the work of Gardner and Collins (Gardner and Collins, 2000) and a more complex discrete stochastic model based on the work of Isaacs (Isaacs, 2003). The tri-stable switch, designed, modelled, and partially constructed as an iGEM 2006 project at Brown University, is to be composed entirely of Biobricked parts from the Registry of Standard Biological Parts. In addition to providing support for the iGEM hypothesis, the tri-stable toggle switch has implications for biotechnology and gene therapy.