Team:IISER-Mohali-INDIA/Improve

Improve

Peshawar team in iGem 2016 attempted to detect the level of CO and NO₂ using a single reporter system. We improved upon the project with an unique circuit that can be used for different pollutants whose promoters are known. Apart from detection we also intend to capture the pollutants and use them by co-culture with a suitable microbial system.

Pollutants in the biosphere pose a major threat to most organisms. Our project seeks their containment through detection and capture. Current technologies like PTR-MS aren’t cost effective and have limited deployment. Luckily, our practical alternative (gECO) bypasses these limitations, by harnessing the power of millions of years of evolution, coupled with a few months of ingenious genetic engineering (different cassette for different substrates).

Inspired by control systems engineering, gECO utilizes positive feedback loops to increase sensitivity and negative feedback loops to ensure stability, thereby ensuring the detection and capture of various toxic substances. The co-culturing of these engineered E.coli with naturally existing microbes (that can utilize these noxious substances) ensures their consumption. The hardware integration is deceptively simple: paper strips with LB and glycerol (in a particular ratio) house the engineered E.coli. On contact water, the cells activate expressing chromoproteins in proportional measure to the external pollutant concentration. Overall, the setup is lightweight, aesthetic, cost effective, ensures detection, capture and consumption of various toxic substances.

Parts

Composite Part:

Part no. 1

Part name- Plac->RBS->amilCP Blue chromoprotein

Part number- BBa_K2320001

Description:

AmilCP blue chromoprotein has been cloned under IPTG inducible lac promoter and RBS. Induction of E. coli carrying K2320001 with IPTG leads to intracellular production of blue colored amilCP protein. Other teams can clone their gene of interest along with a RBS of their choice downstream of this IPTG inducible reporter. Induction with IPTG will give a real-time crude estimation of expression of their protein of interest. In addition, this construct will give an added advantage to tune the expression of their protein of interest by varying amount of IPTG or glucose.

Part no. 2

Part name- Plac->RBS->amilGFP BBa_K592010 (yellow)

Part number- BBa_K2320002

Description:

AmilGFP yellow chromoprotein has been cloned under IPTG inducible lac promoter and RBS. Induction of E. coli carrying K2320002 with IPTG leads to intracellular production of yellow colored amilGFP protein. Other teams can clone their gene of interest along with a RBS of their choice downstream of this IPTG inducible reporter. Induction with IPTG will give a real-time crude estimation of expression of their protein of interest. In addition, this construct will give an added advantage to tune the expression of their protein of interest by varying amount of IPTG or glucose.

Part no. 3

Part name- Plac->RBS->amilCP Blue chromoprotein

Part number- BBa_K2320001

Description:

AmilCP blue chromoprotein has been cloned under IPTG inducible lac promoter and RBS. Induction of E. coli carrying K2320001 with IPTG leads to intracellular production of blue colored amilCP protein. Other teams can clone their gene of interest along with a RBS of their choice downstream of this IPTG inducible reporter. Induction with IPTG will give a real-time crude estimation of expression of their protein of interest. In addition, this construct will give an added advantage to tune the expression of their protein of interest by varying amount of IPTG or glucose.

Part no. 4

Part name- Plac->RBS->cjBlue green chromoprotein

Part number- BBa_K2320004

Description:

cjBlue green chromoprotein has been cloned under IPTG inducible lac promoter and RBS. Induction of E. coli carrying K2320004 with IPTG leads to intracellular production of greenish-blue colored chromoprotein. Other teams can clone their gene of interest along with a RBS of their choice downstream of this IPTG inducible reporter. Induction with IPTG will give a real-time crude estimation of expression of their protein of interest. In addition, this construct will give an added advantage to tune the expression of their protein of interest by varying amount of IPTG or glucose.

Part no. 5

Part name- Plac->RBS->eforRed chromoprotein

Part number- BBa_K2320005

Description:

eforRed chromoprotein has been cloned under IPTG inducible lac promoter and RBS. Induction of E. coli carrying K2320005 with IPTG leads to intracellular production of eforRed chromoprotein. Other teams can clone their gene of interest along with a RBS of their choice downstream of this IPTG inducible reporter. Induction with IPTG will give a real-time crude estimation of expression of their protein of interest. In addition, this construct will give an added advantage to tune the expression of their protein of interest by varying amount of IPTG or glucose.