Team:Tacoma RAINmakers/Parts

Team:ECUST/Lab/Notebook

Our Biobricks

Parts Overview

The Tacoma RAINMakers have built 2 biological arsenic circuits that use a combination of native E. coli and coral genes to detect arsenic ions in solution. The circuits vary in which reporter they produce, either chromoproteins spisPink (from S. pistillata) or amilCP (from A.millepora). Each circuit consists of a repressor, which is an arsenic regulator known as ArsR. This protein binds to a promoter containing an ArsR binding site (PArsR) and inhibits the transcription of the downstream reporter gene. Only in the presence of a transcription factor, which in our case is arsenic, does the reporter become derepressed and the reporter is expressed. This circuit allows us to control chromoprotein expression using a protein sensitive to and capable of binding with arsenic ions in solution.

The most significant hazard in the RAINmaker project would be the use of chemicals containing arsenic. Pure arsenic is not only toxic to humans, but it can also be highly toxic to aquatic life with lasting environmental and ecological effects. Sigma Aldrich warns that arsenic may also cause cancer. To work more safely, Tacoma RAINmakers used the sodium arsenate and sodium arsenite, which are the inorganic derivatives of arsenic that shows significantly less toxicity than the pure arsenic. When using sodium arsenate/sodium arsenite, Tacoma RAINmakers are engaging in a series of stringent safety practices. First, safety glasses will always be used to protect eyes. For hand protection, our team will wear nitrile gloves that equal or exceed required layer thickness (0.11mm). Contaminated gloves will be disposed of with proper glove removal technique (without touching glove's outer surface). To protect the body, our team will wear lab coats during all arsenic-related experiments. RAINmakers have considered air-purifying respirators, but have deemed this precaution unnecessary with proper lab technique use of a biosafety cabinet. Arsenic waste solutions or powder will not be allowed to enter drains or any location outside the lab.

Submitted Parts

BBa_K2673000 pSB1C3/PcArsR_PArsR-spisPink

Part Type: Composite
Description: pSB1C3/PcArsR_PArsR-spisPink is Tacoma RAINMaker’s arsenic detection circuit. It is a 2-part circuit that combines 2 series of promoter-gene sequences on one plasmid. The first part of the circuit, PcArsR, contains an E. coli-derived constitutive promoter (Pc), Ribosome Binding Site, Arsenic Regulator (ArsR), and Double Terminator. This sequence allows for unregulated expression of ArsR. The second part of the circuit contains an E. coli-derived Arsenic Regulator promoter (PArsR), Ribosome Binding Site, spisPink chromoprotein, and Double Terminator. The PArsR sequence has a binding site for ArsR, which is a repressor protein that blocks expression of the reporter. When Arsenic ions are present, the Arsenic Regulator proteins bind to those ions instead, allowing for spisPink chromoproteins to be produced.
Website: http://parts.igem.org/Part:BBa_K2673000

BBa_K2673001 pSB1C3/PcArsR_PArsR-amilCP

Part Type: Composite
Creator: Tacoma RAINmakers
Description: pSB1C3/PcArsR_PArsR-amilCP is Tacoma RAINMaker’s arsenic detection circuit. It is a 2-part circuit that combines 2 series of promoter-gene sequences on one plasmid. The first part of the circuit, PcArsR, contains an E. coli-derived constitutive promoter (Pc), Ribosome Binding Site, Arsenic Regulator (ArsR), and Double Terminator. This sequence allows for unregulated expression of ArsR. The second part of the circuit contains an E. coli-derived Arsenic Regulator promoter (PArsR), Ribosome Binding Site, amilCP chromoprotein, and Double Terminator. The PArsR sequence has a binding site for ArsR, which is a repressor protein that blocks expression of the reporter. When Arsenic ions are present, the Arsenic Regulator proteins bind to those ions instead, allowing for amilCP chromoproteins to be produced and fluoresce blue.
Website: http://parts.igem.org/Part:BBa_K2673001

BBa_K2673002 pSB1C3/PArsR-spisPink

Part Type: Composite
Creator: Tacoma RAINmakers
Description: pSB1C3/PArsR-spisPink is used to test part of the complete 2-part arsenic circuit. The part contains an E. coli-derived Arsenic Regulator promoter (PArsR), Ribosome Binding Site, spisPink chromoprotein, and Double Terminator. The expression of the spisPink chromoprotein is regulated only if ArsR is present. In that case, ArsR will repress the promoter.
Website: http://parts.igem.org/Part:BBa_K2673002

BBa_K2673003 pSB1C3/PcArsR

Part Type: Composite
Creator: Tacoma RAINmakers
Description: The part contains the Ribosome Binding Site, PcArsR, and Double Terminator. This sequence allows for unregulated expression of ArsR, a regulatory protein.
Website: http://parts.igem.org/Part:BBa_K2673003

BBa_K2673004 pSB1C3/PArsR-amilCP

Part Type: Composite
Creator: Tacoma RAINmakers
Description: pSB1C3/PArsR-amilCP is used to test part of the complete 2-part arsenic circuit. The part contains an E. coli-derived Arsenic Regulator promoter (PArsR), Ribosome Binding Site, spisPink chromoprotein, and Double Terminator. The expression of the amilCP chromoprotein is regulated only if ArsR is present. In that case, ArsR will repress the promoter.
Website: http://parts.igem.org/Part:BBa_K2673004

BBa_K2673005 pSB1C3/T7-amilCP

Part Type: Composite
Creator: Tacoma RAINmakers
Description: AmilCP reporter driven by high expression T7 promoter.
Website: http://parts.igem.org/Part:BBa_K2673005