NSF Award
9560186
*** 9560186 Steffan This Small Business Innovation Research Phase I project involves the development of biosensors for measuring trichloroethylene (TCE) and related chlorinated solvents in groundwater. The project involves creating whole-cell biosensors by fusing reporter genes with chlorinated solvent-induced toluene oxidation genes from Pseudomonas mendocina KR1 and Pseudomonas sp. ENVPC5. A variety of reporter genes will be evaluated, but the work will focus on the use of bioluminescence genes that have been widely used for the creation of biosensors. Initial biosensor development will involve creating transcriptional fusions between the reporter genes and cloned toluene oxidation structural genes. Once created, the fusions will be transferred back to the original host organism via marker exchange. Additional biosensors will be created by cloning the entire regulons of the toluene oxidation pathways into E. coli and inserting the reporter genes so it is expressed in response to TCE. This approach will allow an evaluation of the most appropriate host organisms for use as biosensors. The biosensors will be tested for their sensitivity for measuring TCE in ground water, and the expression of the reporter genes will be compared to expression of the wild-type toluene oxidation genes in response to TCE. The biosensors will ultimately be used for developing on-line sensors for monitoring TCE in groundwater, for developing TCE test kits, and for evaluating the in situ performance of TCE-degrading microorganisms. This Phase I research will result in the development of living TCE biosensors that can be used to construct on-line monitoring devices and test kits. They will also aid in evaluating, monitoring and modeling the performance of TCE-degrading bacteria in situ The development of an inexpensive biosensor system will reduce the cost of routine chemical monitoring, allow on-line monitoring, and increase the cost effectiveness of environmental remediation. ***
