Design

In our project, we view a single recombinase and a pair of its target site as a functional unit for building dynamic switches. With this concept in mind, we designed more sophisticated composite parts such as the measurement devices, gene randomizer, logic gates, and pulse generator.

Measurement devices

To quantify orthogonality and recombination efficiency, we have designed simple measurement devices to characterize our basic parts – the five recombinases (Cre, Dre, VCre, SCre, Vika) and their target sites (LoxP, Rox, Vlox, Slox, Vox). They are a set of fifteen biobricks that have a target sites-flanked transcriptional terminator inserted between a constitutive promoter and a RFP:

Thus, when recombination occurred, the transcriptional terminator will be excised, expressing the RFP. We propose to use this set of device and our protocol for standardized measurement of recombination progression using plate reader.

Pulse generator

Aware of the potential for SSR to mediate genetic engineering in a highly dynamic way, we have designed a novel construct that once initiate, will express multiple genes in separated pulses of transcription and translation.

The functionality of the pulse generator has been simulated in the model page , where we have performed stochastic modeling and used it to predict the behavior of the pulse generator in different conditions. The pulse generator may find applications in metabolic engineering, where one can separate temporally the expression of several genes, thus lowering the metabolic stress of a cell (1).