This year, our project has successfully implemented a modular design for an engineered bacterial platform, leading to the rational design and construction of several core functional modules. These modules collectively enable the bacteria to efficiently degrade specific pollutants, enhance their survival in challenging environments, and incorporate essential biosafety features. A comprehensive characterization was conducted on the key genetic components, including their functionality, efficiency, and regulatory mechanisms. We documented the degradation gene clusters for dibenzofuran and toluene, the resistance proteins for salinity and toxic substance tolerance, and the critical parts of the kill switch and quorum sensing system. With the intention of providing a clear parts overview for future synthetic biology endeavors, we hope this summary will facilitate the adoption and optimization of these well-characterized genetic elements in constructing robust biological systems.
The information below provides a detailed description of our new parts.
Name: dfdA1
Type: Protein coding sequences
Description: Part of dfdA1234BC gene cluster, dibenzofuran 4,4a-dioxygenase large subunit.
Name: dfdA2
Type:Protein coding sequences
Description:Part of dfdA1234BC gene cluster, dibenzofuran 4,4a-dioxygenase small subunit.
Name: dfdA3
Type:Protein coding sequences
Description:Part of dfdA1234BC gene cluster, dibenzofuran 4,4a-dioxygenase ferredoxin component.
Name: dfdA4
Type: Protein coding sequences
Description:Part of dfdA1234BC gene cluster, dibenzofuran 4,4a-dioxygenase reductase component.
Name: dfdB
Type: Protein coding sequences
Description: Part of dfdA1234BC gene cluster, the product expressed by this gene is a hypothetical protein.
Name: dfdC
Type: Protein coding sequences
Description:Part of dfdA1234BC gene cluster, the product expressed by this gene is a
putative hydrolase.
Name: xylA
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster, the product expressed by this gene is electron transfer component of xylene monooxygenase.
Name: xylB
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster, the product expressed by this gene is benzyl alcohol dehydrogenase.
Name: xylC
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster, the product expressed by this gene is benzaldehyde dehydrogenase.
Name: xylM
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster, the product expressed by this gene is hydroxylase component of xylene monooxygenase.
Name: xylN
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster, the product expressed by this gene is XylN outer membrane protein.
Name: xylU
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster. The TOL plasmid pWW0 xylN gene product from Pseudomonas putida is involved in m-xylene uptake.
Name: xylW
Type: Protein coding sequences
Description:Part of xylABCMNUW gene cluster, the product expressed by this gene is benzyl alcohol dehydrogenase II.
Name: NhaA
Type: Protein coding sequences
Description: NhaA is located in the inner membrane and functions as an antiporter of Na⁺ and H⁺.
Name: AcrA
Type: Protein coding sequences
Description: AcrAB-TolC multidrug efflux pump subunit AcrA. Acridine resistance protein A.
Name: AcrB
Type: Protein coding sequences
Description:AcrAB-TolC multidrug efflux pump subunit AcrA. Acridine resistance protein A.
Name: TolC
Type: Protein coding sequences
Description:Outer membrane channel, which is required for the function of several efflux systems.
Name: LAA-LAA
Type: Degradation tag (Regulatory)
Description: A synthetic 6-amino acid ClpX-dependent degradation tag engineered for enhanced in vivo degradation efficiency and dynamic synthetic circuit regulation.
Name: NhaA-SsrA
Type: Protein coding sequences
Description: A degradation tag SsrA was added at the end of the NhaA coding sequence, so that it would be degraded by the cells after being expressed in E. coli.
Name: NhaA-LAA-LAA
Type: Protein coding sequences
Description: A degradation tag LAA-LAA was added at the end of the NhaA coding sequence, so that it would be degraded by the cells after being expressed in E. coli.
Name: Doc-LAA-LAA
Type: Protein coding sequences
Description: A degradation tag LAA-LAA was added at the end of the Doc coding sequence, so that it would be degraded by the cells after being expressed in E. coli.
Name: Plac-dfdA1234
Type: Composite part
Description: This part consist of plac (BBa_R0010) and dibenzofuran degradation gene cluster dfdA1234(BBa_25JNJNZG, BBa_25F54YSJ, BBa_25SF45XE, BBa_2540XQ2R). The genes required for the degradation of toluene are clustered together in a multi-antisense form and placed after plac. Each coding sequence is preceded by an RBS (BBa_B0034) sequence, and the terminator is located at the end.
Name: Plac-dfdBC
Type: Composite part
Description: This part consist of plac (BBa_R0010) and dibenzofuran degradation gene cluster dfdBC (BBa_25AZV9LK, BBa_25YNB7FF). The genes required for the degradation of toluene are clustered together in a multi-antisense form and placed after plac. Each coding sequence is preceded by an RBS (BBa_B0034) sequence, and the terminator is located at the end.
Name: Plac-xylABCMNUW
Type: Composite part
Description: This part consist of plac (BBa_R0010) and dibenzofuran degradation gene cluster xylABCMNUW (BBa_25WYGEWP, BBa_251TTV02, BBa_25L7CY1E, BBa_25BIZEPA, BBa_25N0E50R, BBa_258960EO, BBa_25W79JYQ). The genes required for the degradation of toluene are clustered together in a multi-antisense form and placed after plac. Each coding sequence is preceded by an RBS (BBa_B0034) sequence, and the terminator is located at the end.
Name: QS-LuxRm-pluxR-LuxI
Type: Composite part
Description: The system is used to detect and express upstream signals with a positive feedback pLuxR-LuxI.