Design

Since the current solution to the blockage is not environmentally friendly and economical, we hope to use synthetic biology to solve the three problems of rust deposition, biofilm accumulation and microbial activity by engineering microbe.

However, the three functions of engineered bacteria may bring great metabolic pressure to the engineered bacteria, and the anti-bacterial substances produced by them may adversely affect their growth and reproduction. So we need to design a set of genetic circuit to improve this situation and achieve a sequential expression of these three functions!

Inspired by the way people currently deal with it, people first add rust remover and biofilm remover, which first destroy the living environment of the microorganisms, causing them to fail to receive protection, and then bactericide is added to achieve high bactericidal effect. Therefore, in our gene circuit, it is necessary to perform the function of destroying the microenvironment of the microbial before performing the sterilization function.

To perform the function of destroying the microenvironment, the very first thing is to sense it!

Based on the principles of quorum sensing, we realized the identification of bacterial microenvironment. This enables our engineered E. coli to recognize the quorum sensing signal molecule AHL produced by iron bacteria, activate the expression of biological rust removers and biological biofilm remover. The biological rust remover we chose is the strongest ferric chelating agent-siderophore in nature. As for biofilm remover we chose a glycosidase from the actinomycetemella actinomycete that degrades the biofilm polysaccharide backbone.

With the progress of rust removing, the concentration of siderophore-Fe³⁺ complex will continue to increase, and we hope that after the bacteria sense this signal, the expression of the biocide will be initiated. But in the natural iron sensing system, high concentration of iron inhibits the expression of genes, so we need to modify them. By introducing the foreign gene - lacI, which was inhibited under high-iron conditions and will inhibit the biocide gene, we can realize the design of the iron reversal system.

The biocides we tried to use is cecropin AD, an antibacterial peptide derived from silkworm, which has a spectral and efficient bactericidal effect. In order to release this peptide to extracellular space, we achieve cell rupture by simultaneously expressing the autolysin gene.

For biosafety reasons, we should try to prevent engineered bacteria from escaping into the natural environment from pipeline systems. Given the dark environment inside the pipeline and the light environment is natural space, the light-on suicide is adopted, and the suicide gene is expressed when light is present.

For higher biosafety and detectability, we have designed a new hardware - iTube that can be applied to the pipeline system. iTube's built-in sensors can detect circulating water parameters, then by using artificial neural networks to conduct corrosion prediction and blockage assessment of cooling water systems, it can people to add engineered bacteria for cleaning the biofilm and rust. At the same time, the built-in light system can also be manually turned on and off, making the growth of the engineered bacteria in pipe artificially regulated.

Reference

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