The goal of this project is to engineer a biological circuit that utilizes E. coli plasmid DNA sequences and a green fluorescent protein (GFP) reporter to detect arsenic. The constitutive promoter in our construct produces a regulatory protein ArsR that binds with arsenic ions and triggers visible fluorescence. Our team will demonstrate this circuit using a cell-free lateral flow paper-based system. Ultimately, the purpose of this circuit will be to provide an inexpensive and ubiquitous arsenic sensor for testing environmental samples.
Design and create a synthetic biological circuit containing a reporter protein regulated by arsenic ions.
Demonstrate that the circuit can detect arsenic in solution in cells or using cell-free lysates.
Determine the conditions needed for cell-free lysates to detect EPA limits for harmful concentrations of arsenic.
why we chose arsenic
Arsenic was selected by Cadets2Vets as the first chemical we would attempt to detect using a biological circuit. It is naturally found in the environment, especially in groundwater, and around industrial plants as a byproduct of manufacturing practices.
The detection of arsenic is especially important to team members in the Puget Sound area, because North Tacoma is a Superfund Site. The ASARCO smelting company smokestack, erected in 1917, was a symbol of economic prosperity and booming business in the developing city of Tacoma. The copper smelting site on Ruston’s shore in Puget Sound brought jobs and wealth to the area. Unfortunately, over the course of its 100 years of operation, the plant had been pumping toxic materials like arsenic and lead directly into the air, where it then settled into the surrounding soil and water of the greater Tacoma area.
The smokestack was demolished in 1985, but there are still toxic levels of heavy metals in the soil over 30 years later. The city of Tacoma takes measures to warn new Tacoma homeowners about how to mitigate any increased levels of exposure to toxic materials in their soil, but must also take more drastic steps to protect users of publicly-owned land.
The Department of Defense has a strong interest in developing rapid assays to test for environmental risks to protect the health of service members. Troops often deploy to hostile environments and assessing the risks of the environment itself can be challenging. The development of the arsenic circuit represents a new way to use synthetic biology in the military setting. Demonstrating that a biological circuit can be used in this application will lead to an expansion of the technology to test for more substances in the future.
arsenic levels of interest
The maximum contaminant level for arsenic that federal and state agencies use as guideline to test the arsenic levels in drinking water was established by the EPA. Initially it was set to 50 micrograms/liter or 50 ppb. However, drinking water that contains 50 ppb arsenic for prolonged periods of time may cause bladder or lung cancer and other chronic conditions.
In 2001, the EPA reduced the maximum contaminant level of arsenic to 10 micrograms/ liter or 10 ppb1. Nevertheless, according to data gathered between the years of 1993 and 2007, arsenic contamination of the waters in Washington state persisted2.
Probes in the waters of western part of Washington state, near the southern part of the state by the Columbia River, and also north near Everett have detected higher concentrations of arsenic, close to 50 ppb.
Developing an inexpensive paper-based circuit to detect the concentration of arsenic in the water in the region will greatly influence our knowledge and awareness of the quality of the water are consuming.We hope to develop our assay to detect arsenic within the EPA limit.
“Arsenic Detection in Washington State Public Water Supplies”, doh.wa.gov. http://www.doh.wa.gov/CommunityandEnvironment/DrinkingWater/Contaminants/ArsenicinDrinkingWater/MapofArsenicDetections. Web. Sept 06, 2017.