Synthetic gene regulatory networks for single-stri.. (SynthStripe)
Synthetic gene regulatory networks for single-stripe gene expression
Date du début: 1 août 2012,
Date de fin: 31 juil. 2014
A pivotal question in developmental biology is how cellular gene regulatory networks can respond to a signalling molecule in a concentration-dependent manner. This can now be studied using the modern tools of synthetic biology, an emerging research field that applies engineering approaches to biological systems.This project aims to adapt and develop a band-forming model in E. coli. It will explore systematically the parameter and network space that regulates the formation of a single stripe of gene expression in a morphogen gradient. The full design space of 3-gene networks, where one gene is activated by the morphogen, is 9,710 (isometric networks removed). We are therefore building a flexible network 'scaffold' that will allow the construction of any such circuit and, in particular, the six core topologies predicted in silico to form single stripes.Rational design of gene regulatory networks in vivo is extremely challenging, due to the complex interactions in living cells. Therefore, we are exploring the parameter space that leads to functional band-forming networks by building combinatorial libraries, followed by selection for an appropriate survival pressure.The synthetic networks are engineered with the potential to react as concentration band filters, with respect to a morphogen concentration. The RNA polymerases from the T7 and SP6 phages are used as activators and artificial zinc finger DNA-binding domains, cI and lacI as repressors. The output of the gene regulatory networks is a selectable gene, fused to a green fluorescent protein, which can be used for selection, counterselection and quantification.
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