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In the experiments of Gardner et al., (35) the Escherichia coli in which were injected the engineered plasmid containing both promoters replicated as the experiment was carried through. The set of chemical reactions above does not take this or other complications into account; rather the reactions put forth are a simple set of ingredients required to obtain bistable steady state as well as switching between such states.
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The dwell times were obtained by determining levels of protein A and B at time intervals of length T in the time traces. If protein A had the higher level, then a counter is set to 1; otherwise it is 0. A switch is indicated by a change in the counter in the next interval. For sufficiently large T, there is a range over which the dwell times are independent of T. If T is too large, short transitions are missed. If T is too small, fluctuations are picked up as switches between steady states. It is because of the difficulty in defining what is a true transition in the presence of the rare raggedy switches that we define a switch through the simple algorithm above. Since we are really interested in comparing the dwells in MaxCal and Gillespie traces, to avoid bias, we use the same algorithm throughout with T equal to 1000 Gillespie or MaxCal steps, a step being defined by a change in particle number of A or B. We also verified our distribution of dwell times in different ways, for example by averaging levels of A and B within the interval T and using this to determine whether our counter variable should be set to 0 or 1. Different specialized methods for computing dwell times for toggle switches are also available in the literature. See: Allen, R. J.; Warren, P. B.;; ten Wolde, P. R. Phys. Rev. Lett. 2005, 94, 018104.
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