Team:Hong Kong HKUST/Overview

HKUST iGEM Team 2017

Project Overview

In synthetic biology, while safety is often one of the largest concerns to the field as a whole, there is a lack of standardized designs that can easily be added to a construct without affecting its function. Our project seeks to remedy this situation by creating a standardized safety switch which allows synthetic biologists to easily and effectively knock out a specific gene of interest when it is no longer required.

Our project could potentially have potential uses in both single and multicellular organisms, and for different applications - including but not limited to bioremediation, therapeutics and biosensors.

Our construct consists of three modules:

  • A Sensing module to receive and amplify a signal
  • A Time Control module to give the Sensing module time to amplify the signal before knocking out the gene of interest
  • A Recombination module to produce a recombinase which will separate the gene of interest from the origin of replication
    • Fig. 1 Domilox Design Overview

    The first module in our construct is the Sensing module. The purpose of this module is to respond to the concentration of AHL in the cell and amplify the amount of AHL as a signal for further modules of the construct by using a positive feedback loop which amplifies an initial concentration of AHL and allows for quorum sensing because AHL can diffuse through the cell membrane into other cells.

    The second module in our construct is the Time Control module. The purpose of this module is to produce a time delay in responding to the AHL signal inside the cell and to control the translation of two repressors downstream the sensing promoter in order to turn on the expression of Cre recombinase, which is necessary for the recombination reactions in the Recombination module. In transducing the signal through the Time Control module instead of directly passing it to the Recombination module, we seek to allow the Sensing module sufficient time to amplify the AHL signal with its positive feedback loop which would allow for more effective quorum sensing when the AHL diffuses into surrounding cells.

    The final module in our construct is the Recombination module. In this module, we use Cre recombinase to separate the origin of replication from the rest of the plasmid, in which the gene of interest to be knocked out is contained. This would prevent plasmids which have recombined from being replicated, and would eventually lead to the dilution of the gene of interest in the population.

    Fig. 2 The Effect of Recombination module